PharmacologyBiochemistryandBehavior,Vol.43, pp. 395-403, 1992
0091-3057/92$5.00 + .00 Copyright©1992PergamonPress Ltd.
Printedin the U.S.A. All rightsreserved.
Cocaine and Regional Brain Monoamines in Mice M. G. H A D F I E L D ~ A N D C. M I L I O
Neurochemistry Research Laboratory, Division o f Neuropathology, Department o f Pathology, Medical College o f Virginia Virginia Commonwealth University, Richmond, VA 23298 Received 11 N o v e m b e r 1991 HADFIELD, M. G. AND C. MILIO. Cocaine and regional brain monoamines in mice. PHARMACOL BIOCHEM BEHAV 43(2) 395-403, 1992.-Cocaine HC1 (0, 10, or 50 mg/kg) was injected into adult male ICR mice IP. Thirty minutes later, brains were removed and nine regions were isolated: olfactory bulbs (OB), olfactory tubercles (OT), prefrontal cortex (PC), septum (SP), striatum (ST), amygdala (AMY), hypothalamus (HT), hippocampus (HC), and thalamus (TH). Using high-performance fiquid chromatography, concentrations of norepinephrine (NE), dopamine (DA), serotonin (5-HT), and their major metabolites were determined. At 10 mg/kg cocaine, NE levels were increased in the AMY and its metabolite, 3-methoxy-4-hydroxyphenylglycol(MHPG), was decreased in the PC, AMY, and HT. DA levels were also increased in the AMY, while its intracellular metabolite, dihydroxyphenylacetic acid (DOPAC), was decreased in the ST and its extracellular metabolite, homovanillic acid (HVA), was decreased in the PC. 3-Methoxytyramine (3-MT) levels were not altered in any tissue. 5-HT levels were increased in the AMY, HT, and TH, while its metabolite 5-hydroxyindoleaceticacid (5-HIAA) was decreased in the OB and ST. MHPG/NE ratios were decreased in the PC, AMY, and HT as were those for I-IVA/DA. DOPAC/DA ratios were decreased in the ST and AMY and increased in the SP while those for 3-MT/DA were decreased in the TH and increased in the PC. 5-HIAA/5-HT ratios were decreased in the AMY, HC and TH. At 50 mg/kg cocaine, there was an increase in DA in the TH. There was a decrease in DOPAC, HVA, and 3-MT, as well as the DOPAC/DA ratio in the ST. In the OT, there was a decrease in DOPAC, the DOPAC/DA ratio, 3-MT, and the 3-MT/DA ratio. I-IVAwas increased in the TH. There was a decrease in 5-HIAA in the ST, HT, and TH. The 5-HIAA/5-HT ratio was decreased in the OT, PC, ST, AMY, HT, and TH. Cocaine Mouse
Monoamines HPLC
Norepinephrine
Dopamine
COCAINE'S CNS effects are generally attributed to its action as a potent uptake inhibitor of norepinephrine (NE) (24), dopamine (DA) (14,15) and 5-hydroxytryptamine (5-HT) (23) at synaptic membranes. Cocaine also alters the concentration, release, synthesis, turnover, metabolism, and receptor sensitivity of monoamine neurotransmitters (1,3,6,8,9,11,15,17, 19,21). Yet, we know relatively little about cocaine's regional effects on brain monoamines. In the present study, we analyzed the concentration of NE, DA, 5-HT and their major metabolites in nine strategic mouse brain regions. The metabolite/neurotransmitter ratios were calculated as an indicator of neurotransmitter utilization.
Serotonin
Metabolites
Brain regions
were kept under controlled conditions of temperature and humidity and received standard lab chow and water ad lib. Cocaine was administered IP at a dose of 10 mg/kg to 10 animals. Ten drug-free control animals received physiological saline IP. The experiment was repeated using a dose of 50 mg/kg cocaine. Thirty minutes later, brains were rapidly removed and frozen in liquid nitrogen ( - 3 2 0 ° C ) to isolate the olfactory bulbs (OB) (10 mg/kg only), olfactory tubercles (OT), prefrontal cortex (PC), septum (SP), striatum (ST), amygdala (AMY), hypothalamus (HT), hippocampus (HC), and thalamus (TH). The amount of tissue obtainable from the nucleus accumbens (NA) was inadequate for analysis. The tissues were then weighed and homogenized in sodium acetate buffer that contained isoproterenol (IP) as an internal standard. The homogenate was filtered and centrifuged, and the monoamines were extracted in ascorbate oxidase before injection into our high-performance liquid chromatography (HPLC) system (12,13). The data were recorded as nanograms of monoamine/gram brain tissue for: NE, 3-methoxy-4-hy-
METHOD Adult male ICR mice (5-7 weeks of age) were used exclusively in these experiments (Dominion Laboratories, Dublin, VA). They were housed in an approved animal facility under the supervision of a doctor of veterinary medicine. Animals
i Requests for reprints should be addressed to M. Gary Hadfield, M. D., Box 17, MCV Station, Richmond, VA 23298. 395
A 10 m g / k g NE MHPG MHPG/NE DA DOPAC DOPAC/DA HVA HVA/DA 3-MT 3-MT/DA SER 5-HIAA 5-HIAA/SER B. 50 m g / k g NE MHPG MHPG/NE DA DOPAC DOPAC/DA HVA HVA/DA 3-MT 3-MT/DA SER 5-HIAA 5-HIAA/SER
53 ± 14± 0 22 ± 116 ± 16 ± 0 14 ± 24 ± 0.26 ± 6 ± 0 046 ± 84 ± 40 ± 0 4 5 .4-
C
TABLE
1
72 19 0 029 14 2 0 017 54 0.054 1 0 006 9.8 6 0042
OB
51 11 0.20 107 14 0 14 27 0 29 10 0 081 76 26 036
± ± ± ± ± ± ± ± ± ± ± ± .4-
E
80 15 0.030 14 2 0.022 5 1 0.047 3 0.023 12 2.8* 0038
30 16 0.048 207 15 0 006 15 0 005
40±42 19 ± 2 9 0 44 ± 0 050 1,482 ± 99 68 ± 5.3~t 0 047 ± 0 003~t 123 .4- 7 2 0 081 ± 0 003 60 ± 591" 0 037 .4- 0.004:~ 252 ± 19 90 ± I1 0 37 ± 0 031~"
± ± ± ± .4± ± .4-
42±55 20 ± 3 3 0.48 ± 0.044 1,244 ± 69 101 ± 5 5 0.078 ± 0 003 141 ± 17 0 127 ± 0 026 106 ± 11 0 085 ± 0 009 209 ± 14 112 ± 72 0.52 ± 0 017
24 10 0.35 1,779 51 0.032 108 0 058
48 ± 6 4
± 23 ±06 ± 0.036 ± 158 .4- 15 ± 0 007 ± 15 ± 0 008
E
39 ± 5 3
23 7 0 33 1,399 48 0 037 93 0 071
C
OT
.4± + ± ± ± ± ± ± ± ± ± ± 11 0.7 0.025 13
44 14 0.022 29 1 0 06 3.3 0.17 47 0 072 9.3 2 1 0.031
76 ± 5 4 26 .4- 1 1 0 36 ± 0 028
76± 13 ± 0 20 ± 21 ±
59 21 0.42 26 48 0 26 31 13 12 0 33 109 28 027
C
PC
± ± .4.4-
17 1 6* 0.03 3.7
96 ± 12 21 ± 2 4 0 21 .4- 0.023~?
110 17 0.17 29
58 ± 28 16±2.0" 0 26 ± 0 022¢ 25 ± 2.1 4 5 ± 0.6 0.18 ± 0.027 13 ± 4.61" 0 65 ± 0.20* 17 ± 2.4 0 67 ± 0.096* 116 ± 5.3 27 ± 1.9 023 ± 0015
E
198 ± 26 31 ± 6 7
39 + 5 5
34 + 6.7
209 ± 19 32 ± 9.0
32 ± 3 9
66 ± 14
11
139 .4- 26
129 ± 28
189±
67 ± 8.9§
35 ± 4 9
235 .4-20
62 ± 14
120 4- 8
E
51 ± 3 4
116 ± 12
C
SP C
± ± ± ± ± ± ± ± + ±
100 10 0.002 38 0013 8 3 0 003 10 48 006
7,119 209 0.031 689 0.099 331 0 046 156 132 0 89
± 688 ± 14 4- 0 0 0 1 ± 71 ± 0011 ± 42 ±0004 ± 12 ±68 .4- 0 0 4 9
53 . 4 . 8 1
2,998 211 0.073 547 0 175 235 0 078 99 58 06
41 ± 1 6
E F F E C T S O F C O C A I N E (10 A N D 50 m g / k g ) ON R E G I O N A L B R A I N L E V E L S O F NE, DA, 5-HT, A N D T H E I R M A J O R M E T A B O L I T E S
ST
5,670 121 0 022 483 0 083 219 0040 139 96 0 69
3,166 162 0 048 491 0 161 251 0.078 94 43 05
233 15§ 0 003~: 27 0008 28 0 003 66 44* 004
509 15:~ 0 001~t 61" 0.005 25* 0.004 13 67I" 0 0391"
conttnued
± •4± ± 44± ± ± 4-
±62
± .4± ± ± + ± ± ± ±
43.4-26
E
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< < <
0091-3057/92$5.00 + .00 Copyright©1992PergamonPress Ltd.
Printedin the U.S.A. All rightsreserved.
Cocaine and Regional Brain Monoamines in Mice M. G. H A D F I E L D ~ A N D C. M I L I O
Neurochemistry Research Laboratory, Division o f Neuropathology, Department o f Pathology, Medical College o f Virginia Virginia Commonwealth University, Richmond, VA 23298 Received 11 N o v e m b e r 1991 HADFIELD, M. G. AND C. MILIO. Cocaine and regional brain monoamines in mice. PHARMACOL BIOCHEM BEHAV 43(2) 395-403, 1992.-Cocaine HC1 (0, 10, or 50 mg/kg) was injected into adult male ICR mice IP. Thirty minutes later, brains were removed and nine regions were isolated: olfactory bulbs (OB), olfactory tubercles (OT), prefrontal cortex (PC), septum (SP), striatum (ST), amygdala (AMY), hypothalamus (HT), hippocampus (HC), and thalamus (TH). Using high-performance fiquid chromatography, concentrations of norepinephrine (NE), dopamine (DA), serotonin (5-HT), and their major metabolites were determined. At 10 mg/kg cocaine, NE levels were increased in the AMY and its metabolite, 3-methoxy-4-hydroxyphenylglycol(MHPG), was decreased in the PC, AMY, and HT. DA levels were also increased in the AMY, while its intracellular metabolite, dihydroxyphenylacetic acid (DOPAC), was decreased in the ST and its extracellular metabolite, homovanillic acid (HVA), was decreased in the PC. 3-Methoxytyramine (3-MT) levels were not altered in any tissue. 5-HT levels were increased in the AMY, HT, and TH, while its metabolite 5-hydroxyindoleaceticacid (5-HIAA) was decreased in the OB and ST. MHPG/NE ratios were decreased in the PC, AMY, and HT as were those for I-IVA/DA. DOPAC/DA ratios were decreased in the ST and AMY and increased in the SP while those for 3-MT/DA were decreased in the TH and increased in the PC. 5-HIAA/5-HT ratios were decreased in the AMY, HC and TH. At 50 mg/kg cocaine, there was an increase in DA in the TH. There was a decrease in DOPAC, HVA, and 3-MT, as well as the DOPAC/DA ratio in the ST. In the OT, there was a decrease in DOPAC, the DOPAC/DA ratio, 3-MT, and the 3-MT/DA ratio. I-IVAwas increased in the TH. There was a decrease in 5-HIAA in the ST, HT, and TH. The 5-HIAA/5-HT ratio was decreased in the OT, PC, ST, AMY, HT, and TH. Cocaine Mouse
Monoamines HPLC
Norepinephrine
Dopamine
COCAINE'S CNS effects are generally attributed to its action as a potent uptake inhibitor of norepinephrine (NE) (24), dopamine (DA) (14,15) and 5-hydroxytryptamine (5-HT) (23) at synaptic membranes. Cocaine also alters the concentration, release, synthesis, turnover, metabolism, and receptor sensitivity of monoamine neurotransmitters (1,3,6,8,9,11,15,17, 19,21). Yet, we know relatively little about cocaine's regional effects on brain monoamines. In the present study, we analyzed the concentration of NE, DA, 5-HT and their major metabolites in nine strategic mouse brain regions. The metabolite/neurotransmitter ratios were calculated as an indicator of neurotransmitter utilization.
Serotonin
Metabolites
Brain regions
were kept under controlled conditions of temperature and humidity and received standard lab chow and water ad lib. Cocaine was administered IP at a dose of 10 mg/kg to 10 animals. Ten drug-free control animals received physiological saline IP. The experiment was repeated using a dose of 50 mg/kg cocaine. Thirty minutes later, brains were rapidly removed and frozen in liquid nitrogen ( - 3 2 0 ° C ) to isolate the olfactory bulbs (OB) (10 mg/kg only), olfactory tubercles (OT), prefrontal cortex (PC), septum (SP), striatum (ST), amygdala (AMY), hypothalamus (HT), hippocampus (HC), and thalamus (TH). The amount of tissue obtainable from the nucleus accumbens (NA) was inadequate for analysis. The tissues were then weighed and homogenized in sodium acetate buffer that contained isoproterenol (IP) as an internal standard. The homogenate was filtered and centrifuged, and the monoamines were extracted in ascorbate oxidase before injection into our high-performance liquid chromatography (HPLC) system (12,13). The data were recorded as nanograms of monoamine/gram brain tissue for: NE, 3-methoxy-4-hy-
METHOD Adult male ICR mice (5-7 weeks of age) were used exclusively in these experiments (Dominion Laboratories, Dublin, VA). They were housed in an approved animal facility under the supervision of a doctor of veterinary medicine. Animals
i Requests for reprints should be addressed to M. Gary Hadfield, M. D., Box 17, MCV Station, Richmond, VA 23298. 395
A 10 m g / k g NE MHPG MHPG/NE DA DOPAC DOPAC/DA HVA HVA/DA 3-MT 3-MT/DA SER 5-HIAA 5-HIAA/SER B. 50 m g / k g NE MHPG MHPG/NE DA DOPAC DOPAC/DA HVA HVA/DA 3-MT 3-MT/DA SER 5-HIAA 5-HIAA/SER
53 ± 14± 0 22 ± 116 ± 16 ± 0 14 ± 24 ± 0.26 ± 6 ± 0 046 ± 84 ± 40 ± 0 4 5 .4-
C
TABLE
1
72 19 0 029 14 2 0 017 54 0.054 1 0 006 9.8 6 0042
OB
51 11 0.20 107 14 0 14 27 0 29 10 0 081 76 26 036
± ± ± ± ± ± ± ± ± ± ± ± .4-
E
80 15 0.030 14 2 0.022 5 1 0.047 3 0.023 12 2.8* 0038
30 16 0.048 207 15 0 006 15 0 005
40±42 19 ± 2 9 0 44 ± 0 050 1,482 ± 99 68 ± 5.3~t 0 047 ± 0 003~t 123 .4- 7 2 0 081 ± 0 003 60 ± 591" 0 037 .4- 0.004:~ 252 ± 19 90 ± I1 0 37 ± 0 031~"
± ± ± ± .4± ± .4-
42±55 20 ± 3 3 0.48 ± 0.044 1,244 ± 69 101 ± 5 5 0.078 ± 0 003 141 ± 17 0 127 ± 0 026 106 ± 11 0 085 ± 0 009 209 ± 14 112 ± 72 0.52 ± 0 017
24 10 0.35 1,779 51 0.032 108 0 058
48 ± 6 4
± 23 ±06 ± 0.036 ± 158 .4- 15 ± 0 007 ± 15 ± 0 008
E
39 ± 5 3
23 7 0 33 1,399 48 0 037 93 0 071
C
OT
.4± + ± ± ± ± ± ± ± ± ± ± 11 0.7 0.025 13
44 14 0.022 29 1 0 06 3.3 0.17 47 0 072 9.3 2 1 0.031
76 ± 5 4 26 .4- 1 1 0 36 ± 0 028
76± 13 ± 0 20 ± 21 ±
59 21 0.42 26 48 0 26 31 13 12 0 33 109 28 027
C
PC
± ± .4.4-
17 1 6* 0.03 3.7
96 ± 12 21 ± 2 4 0 21 .4- 0.023~?
110 17 0.17 29
58 ± 28 16±2.0" 0 26 ± 0 022¢ 25 ± 2.1 4 5 ± 0.6 0.18 ± 0.027 13 ± 4.61" 0 65 ± 0.20* 17 ± 2.4 0 67 ± 0.096* 116 ± 5.3 27 ± 1.9 023 ± 0015
E
198 ± 26 31 ± 6 7
39 + 5 5
34 + 6.7
209 ± 19 32 ± 9.0
32 ± 3 9
66 ± 14
11
139 .4- 26
129 ± 28
189±
67 ± 8.9§
35 ± 4 9
235 .4-20
62 ± 14
120 4- 8
E
51 ± 3 4
116 ± 12
C
SP C
± ± ± ± ± ± ± ± + ±
100 10 0.002 38 0013 8 3 0 003 10 48 006
7,119 209 0.031 689 0.099 331 0 046 156 132 0 89
± 688 ± 14 4- 0 0 0 1 ± 71 ± 0011 ± 42 ±0004 ± 12 ±68 .4- 0 0 4 9
53 . 4 . 8 1
2,998 211 0.073 547 0 175 235 0 078 99 58 06
41 ± 1 6
E F F E C T S O F C O C A I N E (10 A N D 50 m g / k g ) ON R E G I O N A L B R A I N L E V E L S O F NE, DA, 5-HT, A N D T H E I R M A J O R M E T A B O L I T E S
ST
5,670 121 0 022 483 0 083 219 0040 139 96 0 69
3,166 162 0 048 491 0 161 251 0.078 94 43 05
233 15§ 0 003~: 27 0008 28 0 003 66 44* 004
509 15:~ 0 001~t 61" 0.005 25* 0.004 13 67I" 0 0391"
conttnued
± •4± ± 44± ± ± 4-
±62
± .4± ± ± + ± ± ± ±
43.4-26
E
*p §p "l'P ~p
< < <
