I. MOOD STABILIZERS ANTIINFLAMMATORY EFFECT OF LITHIUM CHRONIC LITHIUM REDUCES INFLAMMATION-RELATED UPREGULATED BRAIN ARACHIDONIC ACID METABOLISM IN HIV-1 TRANSGENIC RAT. HIV-1 transgenic (Tg) rats, a model for human HIV-1 associated neurocognitive disorder (HAND), show upregulated markers of brain arachidonic acid (AA) metabolism and neuroinflammation after 7 months of age. Since lithium decreases brain AA metabolism in control rats and a rat lipopolysaccharide model of neuroinflammation, and appears to do so in HAND, we hypothesized that lithium would dampen upregulated brain AA metabolism in HIV-1 Tg rats. We confirmed this by measuring regional brain AA incorporation using quantitative autoradiography, after intravenous 1-(14)CAA infusion in unanesthetized 10-month-old HIV-1 Tg and age-matched wildtype that had been fed a control or LiCl diet for 6 weeks. Incorporation was significantly higher in HIV-1 Tg than wildtype rats fed the control diet. Since lithium treatment dampened upregulated brain AA metabolism in HIV-1 Tg rats, it may prove therapeutic in HIV-1 patients with HAND and neuroinflammation through a comparable effect. (1). TESTING NONTERATOGENIC SUBSTITUTES FOR VALPROATE PROPYLISOPROPYLACETIC ACID (PIA), A CONSTITUTIONAL ISOMER OF VALPROIC ACID, UNCOMPETITIVELY INHIBITS ARACHIDONIC ACID ACYLATION BY RAT ACYL-COA SYNTHETASE 4: A POTENTIAL DRUG FOR BIPOLAR DISORDER Mood stabilizers used for treating bipolar disorder (BD) selectively downregulate arachidonic acid (AA) turnover (deacylation-reacylation) in brain phospholipids, when given chronically to rats. In vitro studies suggest that one of these, valproic acid (VPA), which is teratogenic, reduces AA turnover by inhibiting the brain long-chain acyl-CoA synthetase (Acsl)4 mediated acylation of AA to AA-CoA. We tested whether non-teratogenic VPA analogues might also inhibit Acsl4 catalyzed acylation, and thus have a potential anti-BD action. Rat Acsl4-flag protein was expressed in Escherichia coli, and the ability of three VPA analogues, propylisopropylacetic acid (PIA), propylisopropylacetamide (PID) and N-methyl-2,2,3,3-tetramethylcyclopropanecarboxamide (MTMCD), and of sodium butyrate, to inhibit conversion of AA to AA-CoA by Acsl4 was quantified using Michaelis-Menten kinetics. Acsl4-mediated conversion of AA to AA-CoA in vitro was inhibited uncompetitively by PIA, with a Ki of 11.4mM compared to a published Ki of 25mM for VPA, while PID, MTMCD and sodium butyrate had no inhibitory effect. PIA's ability to inhibit conversion of AA to AA-CoA by Acsl4 in vitro suggests that, like VPA, PIA may reduce AA turnover in brain phospholipids in unanesthetized rats, and if so, may be effective as a non-teratogenic mood stabilizer in BD patients. (2) VALNOCTAMIDE (VCD), SHOWN EFFECTIVE IN BIPOLAR DISORDER, INHIBITS ACSL-4 ACTIVITY IN VITRO. We are finding that VCD, a nonteratogenic analog of valproate, inhibits Acsl-4 activity in vitro like valproic acid. Since VCD was useful in BD patients and inhibits a key enzyme of brain phospholipid AA turnover cycle, we are extending our study to test VCD on AA turnover on our in vivo rat model. HOMOLOGY MODELING OF INHIBITORS OF ACSL-4. In our earlier studies we reported that Acsl-4 was inhibited by valproic acid (VPA) and its analogues PIA and VCD. The mammalian structure of Acsl-4 enzyme is not available. To give a new direction of new drug discovery, we are studying the homology modeling of mammalian Acsl-4. If we succeed, we may able to design new drugs that are less teratogenic and less hepatotoxic mood stabilizers than VPA. II. ATYPICAL ANTIPSYCHOTICS CHRONIC CLOZAPINE DOWNREGULATES MARKERS OF RAT BRAIN ARACHIDONIC ACID CASCADE AND SYNAPTIC INTEGRITY. Clozapine is an atypical antipsychotic used for treating schizophrenia (SZ) and BD. In both diseases brain AA cascade markers are upregulated. Similar to olanzapine, chronic clozapine reduced rat brain AA cascade markers: cyclooxygenase (COX) activity and concentration of proinflammatory PGE2. It also showed neuroprotective action, by increasing brain expression of brain derived neurotrophic factor (BDNF) and of a marker of post-synaptic dendritic spines, drebrin. These effects were associated with increased expression of docosahexaenoic acid (DHA)-selective calcium-independent type VIA iPLA2. These results show overlap with effects of mood stabilizers and olanzapine, and support targeting of the brain AA cascade as a common mechanism of action of both classes of drug. Dietary deprivation of AA and its precursor linoleic acid (LA, 18:2 n-6)could enhance clozapines antipsychotic (3) . CHRONIC CLOZAPINE REDUCES RAT BRAIN ARACHIDONIC ACID METABOLISM BY REDUCING PLASMA ARACHIDONIC ACID AVAILABILITY. Chronic administration of mood stabilizers to rats down-regulates the brain arachidonic acid (AA) cascade. This down-regulation may explain their efficacy against bipolar disorder (BD), in which brain AA cascade markers are elevated. The atypical antipsychotics, olanzapine (OLZ) and clozapine (CLZ), also act against BD. When given to rats, both reduce brain cyclooxygenase activity and prostaglandin E(2) concentration;OLZ also reduces rat plasma unesterified and esterified AA concentrations, and AA incorporation and turnover in brain phospholipid. To test whether CLZ produces similar changes, we used our in vivo fatty acid method in rats given 10 mg/kg/day i.p. CLZ, or vehicle, for 30 days;or 1 day after CLZ washout. 1-(14) CAA was infused intravenously for 5 min, arterial plasma was collected and high-energy microwaved brain was analyzed. CLZ increased incorporation coefficients k * and decreased corrected rates J(in,i) of plasma unesterified AA into brain phospholipids. These effects disappeared after washout. Thus, CLZ and OLZ similarly down-regulated kinetics and cyclooxygenase expression of the brain AA cascade, likely by reducing plasma unesterified AA availability. Atypical antipsychotics and mood stabilizers may be therapeutic in BD by down-regulating, indirectly or directly respectively, the elevated brain AA cascade of that disease. (4). III. ANTIDEPRESSANTS TEMPORARY POSTNATAL FLUOXETINE ADMINISTRATION TO MICE DECREASES BRAIN ARACHIDONIC ACID METABOLISM DURING ADULTHOOD. Fetal or postnatal exposure of mice or humans to the antidepressant fluoxetine alters their later behavior. We are testing whether changes associated with the altered behavior caused by post-natal fluoxetine involve arachidonic acid (AA), a second messenger released following 5-HT2A/2C receptor-mediated activation of brain cytosolic phospholipase A2 (cPLA2).
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