European Journal ~Pharmacology, 215 (1992) 185 189

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(c~ 1992 Elsevier Science Publishers B.V. All rights reserved 0014-2999/92/$115.00

EJP 52422

Effects of i.c.v, lithium chloride administration on m o n o a m i n e concentration in rat mediobasal hypothalamus M a t i i d e E. O t e r o L o s a d a a n d M o d e s t o C. R u b i o hlstituto de lm'estigaciones Farmacol~'>gicas, Consejo Nacional de lncestigaciones Cient[]~cas y T~cnicas (ININFA) (CONICET), Junht 956 50 pO, Buenos Aires 1113, Argentina Received 9 December 1991, revised MS received 6 February 1992, accepted l l February 1992

We investigated the acute effects of a single i.c.v, injection of lithium chloride (LiC1) the neuroamine content of the rat mediobasal hypothalamus (MBH). The effects of lithium on amine synthesis and degradation enzymes were also studied in vitro. Noradrenaline (NA), dopamine (DA), serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) concentrations were reduced 10 rain after i.c.v, injection of 24 nmol of LiCI and returned to control values 30 min after the injection. Two nmol of LiCI reduced the concentration of DA (10 and 30 min after injection) and 5-HIAA (30 rain after injection). LiCI (0.5-10 raM) inhibited tyrosine hydroxylase activity (catecholamine synthesis) in vitro in a concentration dependent manner. The i.c.v. administration of a high dose of LiCI reduced the content of neuroamines in the MBH. This might result from and inhibition of synthesis. A possible link between the observed changes and some reported side effects of lithium therapy is discussed. Lithium chloride; Monoamines; Hypothalamus; (i.c.v.)

1. Introduction

Lithium salts are widely used in the treatment of manic-depressive psychosis (Schou, 1968) and modify neurotransmission in different systems in the brain (Katz et al., 1968; Knapp and Mandell, 1973, 1975; Poitou and Bohuon, 1975; Otero Losada and Rubio, 1986a,b). Neuroendocrine alterations and hypothermia usually a p p e a r as side effects during treatment with lithium and also as signs of toxicity due to high lithium doses. Monoamines in the hypothalamus are involved in the regulation of body t e m p e r a t u r e (Hellon, 1974; Lin et al., 1983; Simpson and Resch, 1986) and also influence the release of hypothalamic hormones, which are reported to be altered by lithium administration in both humans ( G r o f et al., 1984; Joffe et al., 1986; Seely et al., 1989) and animals (Banerji et al., 1982; Mc Cance et al., 1989; Salata and Klein, 1987). Experimental evidence thus points at the hypothalamus as a likely site of action for lithium at least in relation with some side effects observed during treatment.

We have previously observed alterations in the metabolism of monoamines in localized areas of the rat brain after both acute (Otero Losada and Rubio, 1984, 1985) and chronic (Otero Losada and Rubio, 1986a) lithium chloride (LiCI) i.p. administration. These studies did not include the hypothalamus. As far as we know there is no information concerning the possible effects of i.c.v, lithium administration on monoaminergic pathways in the hypothalamus. Furthermore lithium has rarely been given i.c.v. Hence in the present study we investigate the effects of acute i.c.v. LiC1 administration on the content of monoamines in the rat mediobasal hypothalamus (MBH), an area rich in serotonergic and noradrenergic terminals which might participate in the control of body temperature and hypothalamic hormone release. The activities of the enzymes of synthesis and degradation of monoamines were also measured in the presence of LiCI in vitro.

2. Materials and m e t h o d s 2.1. A n i m a l s and treatment

Correspondence to: M.E. Otero Losada, Instituto de Investigaciones Farmacol6gicas, Consejo Nacional de Investigaciones Cient[ficas y T6cnicas (ININFA) (CONICET), Junfn 956 5 o p 0 Buenos Aires 1113, Argentina. Tel. 54.1.961 6784 or 5949.

For i.c.v, administration, a polyethylene cannula was chronically implanted in the third ventricle of male Wistar rats (150-200 g). After 10-15 days of recovery, LiCI, 2 or 24 nmol, (dissolved in a 0.9% NaC1 solution)

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was slowly administered (2 pA/min) as a single i.c.v. injection (2 pA/rat). Control rats received an equal volume of 0.9% NaC1 solution. The rats were decapitated at different times after the i.c.v, injection (10, 30 min) and the mediobasal hypothalami were dissected out (Hompes et al., 1982; Palkovits and Brownstein, 1988) at 0°C and stored at -60°C until high-pressure liquid chromatography (HPLC) determinations or radioenzymatic assays. For determination of Li + concentration, CSF samples were collected through the implanted cannula by means of aspiration with a syringe, deproteinized and read at 684 nm excitation wavelength by atomic emission spectrophotometry. 2.2. HPLC determinations with electrochemical detection (HPLC-EC) The endogenous concentrations of noradrenaline (NA), dopamine (DA), serotonin (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA) were determined by HPLC-EC as described previously (Otero Losada, 1988). Briefly the tissues were homogenized and deproteinized in a 0.4 M HCIO 4 solution containing 0.1% Na2-EDTA and 0.1% Na2SO 3. After centrifugation, aliquots (50 ~I) of the supernatants were analysed by HPLC by means of a liquid chromatoggrapher 5000 Varian with electrochemical detector. Chromatographic conditions were: (a) 5-HT and 5-HIAA detection, mobile phase 0.1 M citric acid, 0.2 M sodium phosphate dibasic, 8% methanol; pH 4.8; flow rate 1 ml/min; electrode potential +0.7 V; (b) NA and DA detection, mobile phase 0.1 M sodium phosphate monobasic, 0.1 mM EDTA, 0.2 mM octyl-sulphonic acid (sodium salt), 4% methanol; pH 4.0; flow rate 1 ml/min; electrode potential +0.7 V. A Micropack MCH-5 (reverse phase) column was used (sensitivity 0.1 pmol; linearity up to 50 pmol).

1974) was assayed in homogenates in the presence of substrate dihydroxybenzoic acid (1 mM) and cofactor (14C)-S-adenosyl-l-methionine (0.1 raM) (saturating concentrations). 2.4. Drugs and stat&tical analys& Tyrosine-L-l-[1-14C], 57.9 Ci/mol (New England Nuclear, NEN); Tyramine-[3H(G)], 14.0 Ci/mmol (NEN); S-adenosyl-L-methionine-[methyl-(14C)], 56.7 mCi/mmol (NEN); 6,7-dimethyltetrahydropteridine (DMPH 4) (Aldrich, A, USA); dopamine (3-hydroxytyramine)clorhydrate, norepinephrine bitartrate, 3,4-dihydroxybenzoic acid (protocatechuic acid) (Sigma); acetonitrile (Sintorgan, Bs As, Argentina). Lithium chloride (Mallinckrodt; New York, USA). Results were analysed with a two-way analysis of variance followed by Tuckey's t-test.

3. Results

Animals injected i.c.v, with LiCI had piloerection and reduced spontaneous motor activity, responses which were more evident in animals that had received the higher LiCI dose. Lithium levels in CSF, based on three to four pooled samples from five to six rats, were (mM) 0.33 _+0.02 and 2.15 _+0.37 (10 min after i.c.v. injection of 2 or 24 nmol of LiC1, respectively), 0.19 +_ 0.03 and 1.34 _+ 0.17 (30 min after i.c.v, injection of 2 or 1.0-

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The assays were performed according to the original methods slightly modified as described previously (Otero Losada and Rubio, 1984, 1989). Tyrosine hydroxylase (TH) activity (Waymire et al., 1971) was measured in tissue slices in the presence of substrate [1-14C]tyrosine 0.01 mM (non-saturating concentration) (58.1 mCi/mmol), or in homogenates in the presence of [1-~4C]tyrosine 0.1 mM (10.5 mCi/mmol) and cofactor 6,7-dimethyltetrahydropteridine (DMPH 4) 0.1 mM (saturating concentrations). Monoamineoxidase (MAO) (Mc Caman et al., 1965) was measured with [3H]tyramine 0.01 mM (nonsaturating concentration) (2 mCi/mmol) as substrate in tissue slices, or with 0.1 mM (saturating concentration) (40 mCi/mmol) in homogenates. Catechol-O-methyl-transferase (COMT) (Jarrott,

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'l~me (,r, in) Fig. l. Effect of LiCI i.c.v, on NA, D A , 5 - H T and 5 - H I A A c o n c e n t r a tion in the m e d i o b a s a l h y p o t h a l a m u s . O r d i n a t e s : / x g / g tissue. Abscissas: time (min) after i.c.v. LiC1 injection, o Control; • 2 nmol L i C k • 124 nmol LiCI. R e s u l t s are e x p r e s s e d as the means_+ S.E. of 6 - 1 0 a n i m a l s p e r group. * P < 0 . 0 5 , * * P < 0 . 0 2 5 , ~ P < 0 . 0 0 5 , ~# P

Effects of i.c.v. lithium chloride administration on monoamine concentration in rat mediobasal hypothalamus.

We investigated the acute effects of a single i.c.v. injection of lithium chloride (LiCl) the neuroamine content of the rat mediobasal hypothalamus (M...
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