°' BRIEF REPORTS Effect of Age, Sex Steroids, Brain Region, and Genetic Strain on Brain Inositol Monophosphatase Activity Yardena Patishi, Robert H. Belmaker, and Galila Agam Key Words: Inositol-monophosphatase, rat brain, lithium, age, sex, hormones BIOL PSYCHIATRY 1996;40:656--659 Introduction Inositol monophosphatase (I-M-P'ase), .which catabolizes inosi- tol monophosphate to inositol (Hallcher and Sherman 1980), is a key enzyme in the phosphatidylinositol (PI) cycle that serves as a second messenger system for several brain neurotransmitters (Belmaker et al 1990). Inhibition of inositol monophosphatase by Li (Ki = 0.86 mmol/L) (Moscovich et al 1990) reduces inositol levels by up to 30% (Sherman et ai 1985; Sherman et al 1987). Inositol reduction has been shown to be physiologically impor- tant, since replacement by exogenous inositol has been shown to reverse Li's effects in several systems, such as Li-pilocarpine seizures in rats (Kofman et al 1992). Li has powerful therapeutic and prophylactic effects on both phases of bipolar manic- depressive illness (Belmaker 1981). Berridge (1989) proposed that Li's reduction of inositol levels would lead to reduced resynthesis of phosphatidylinositol only in overactive systems, thus leading to their effective dampening. The mechanism of Li's inhibition of inositol monophosphatase is uncompetitive (Nahorski et al 1991). Thus, increasing sub- strate concentrations to compensate for Li's reduction of the rate of inositol monophosphatase catabolism is particularly ineffec- tive, since elevated substrate concentrations potentiates the inhibition; however, inositol formation will be directly propor- From the Ministry of Health Mental Health Center (YP, RHB); Laboratory of Biochemistry, Soroka Medical Center (GA); and Unit of Clinical Biochemistry, Faculty of Health Sciences, Ben-Gurion University of the Negev (YP, RHB, GA), Beer Sheva, Israel. Address reprint requests to Galila Agam, PhD, Director, Laboratory of Biochem- istry, Soroka Medical Center, P.O.B. 151, Beer Sheva 84101, Israel. Received August 31, 1995; revised April 23, 1996. tional to the quantity of enzyme protein even in the presence of constant concentrations of Li. Therefore, putative mechanisms that control the activity of inositol monophosphatase may affect the physiological conditions in which Li is more or less effective in reducing inositol synthesis. Materials and Methods The Assay of lnositol Monophosphatase Activity The animals were sacrificed by decapitation. Brains were imme- diately removed, dissected on ice, and kept at -70°C until assayed according to Hallcher and Sherman (1980). All subse- quent steps except the incubation period were performed at 4°C. Enzyme activity was measured in brain homogenates obtained by the addition of 8 mL of homogenization buffer [50 mmol/L Tris-HC1 pH 8.3, 0.2 mol/L KCI, 0.5 mmol/L ethylenediami- netetraacetic acid (EDTA) and 0.1 mmol/L ethylenglycoltet- raacetic acid (EGTA)] to 0.1 g of tissue. Brain homogenization was carried out in a cell disrupter followed by 15 min centrifu- gation at 7500 g at 4°C. The supernatant was used as the source of the enzyme. The reaction mixture for the measurement of inositol monophosphatase activity contained, in a final volume of 210 p~L, 0.7 mmol/L inositol-l-P, 50 mmol/L Tris-HC1 pH 7.8, 250 mmol/L KC1, 3 mmol/L MgC12, and 90 IxL enzyme preparation. Incubation was carried out for 1 hour at 37°C and then 10 p~L of 100% solution trichloroacetic acid (TCA) was added to stop the reaction. The mixture was centrifuged for 15 minutes at 7500 g at 4°C. Inorganic phosphate in the supernatant was determined as described (Taussky et al 1953). To distinguish © 1996 Society of Biological Psychiatry 0006-3223/96/$15.00 PII S0006-3223(96)00267-3