Erythrocyte but not synaptic human acetylcholinesterase hydrolyzes heroin to morphine

S43 proteins, and stimulation of type I, III or VIII adenylyl cyclase by elevating intracellular calcium concentration and the consequent activation of calcium/calmodulin. ODOR DISCRIMINATION LEARNING IS CORRELATED WITH REDUCED AFTER-HYPERPOLARIZATION IN PIRIFORM CORTEX PYRAMIDAL CELLS D. Saar, Y. Grossman and E. Barkai Dept. of Physiology, Faculty of Health Sciences and Zlotowsky Center for Neuroscience, Ben-Gurion University of the Negev, Beer-Sheva, Israel Learning-related cellular modifications were studied in the rat piriform cortex. Water deprived rats were divided into 3 groups: a ‘trained’ group was trained in a 4-arm maze to discriminate positive cues in 35– 50 pairs of odors (‘extensive training’), a ‘control’ group was ‘pseudotrained’ by random water rewarding, and a ‘naive’ group was not exposed to training. Intrinsic properties and synaptic inputs of layer II pyramidal neurons were examined in brain slices by intracellular recordings with sharp electrodes. Neurons from the three animal groups (7 animals in each group) did not differ in their passive membrane properties and single action-potential characteristics. However, the after-hyperpolarizations (AHPs) that follow trains of six action potentials were reduced by 40% (P < 0.01) in neurons of ‘trained’ rats (mean ± SD: 4.4 ± 1.5 mV, n = 10) than in ‘naive’ (6.9 ± 3.0 mV, n = 13) and in ‘control’ groups (7.8 ± 2.8 mV, n = 11), suggesting increased excitability in these cells. A significant, although smaller reduction (20%) in AHPs was observed in pyramidal neurons as soon as rats demonstrated enhancement of learning rate (‘mild training’), usually after being trained to discriminate between one or two pairs of odors (6.6 ± 1.9 mV, n = 28, in ‘trained’, vs. 8.2 ± 1.5 mV, n = 20, in ‘control’ and 8.2 ± 2.7 mV, n = 11, in ‘naive’ group). AHPs were reduced in most of the sampled neurons. AHP remained reduced up to 3 days after the last training session. However, after 5 days of training suspension AHP amplitude increased back to the pre-training value. Accordingly, training suspension for 5 days or more resulted in slower learning of new odors. We suggest that the reduced AHP following olfactory learning is related to the ability of the cortical network to enter a ‘leaning mode’ which creates favorable conditions for a later occurring enhanced synaptic release. Supported by grants from the National Institute for Psychobiology in Israel and the Israeli Ministry for the Arts and Science. TRANSIENT SYNAPTIC MODIFICATIONS IN PIRIFORM CORTEX PYRAMIDAL CELLS ASSOCIATED WITH ODOR LEARNING D. Saar, D. Lebel, Y. Grossman and E. Barkai Dept. of Physiology, Faculty of Health Sciences and Zlotowsky Center for Neuroscience, Ben-Gurion University, Beer-Sheva, Israel Learning-related cellular modifications were studied in the rat piriform cortex. Water deprived rats were divided into 3 groups: a ‘trained’ group was trained in a 4-arm maze to discriminate positive cues in 35– 50 pairs of odors (‘extensive training’), a ‘control’ group was ‘pseudotrained’ by random water rewarding, and a ‘naive’ group was not exposed to training. Synaptic connections between layer II pyramidal neurons were examined in brain slices by extracellular and intracellular recordings. Kinetics of single EPSPs were similar in all groups. However, paired-pulse facilitation (PPF) was significantly smaller in neurons from the ‘extensively trained’ group, at inter-stimulus intervals ranging between 50–150 ms (e.g. at 50 ms: 1.19 ± 0.17, mean ± SD, n = 22, in ‘trained’ group, vs. 1.37 ± 0.25, n = 13, in ‘control’, and 1.32 ± 0.2, n = 20, in ‘naive’ group, P < 0.02). Similar experiments were performed in slices from ‘mildly trained’ rats (after training for 1– 2 pairs of odors). Three days after training completion PPF was reduced in cells from ‘trained’ animals (1.18 ± 0.14, n = 16) as compared to ‘control’ animals (1.33 ± 0.17, n = 17, P < 0.02). The PPF remained

reduced for up to 8 days after training. Further, post-tetanic potentiation (PTP) induced by 2 min repetitive theta stimulation at 50 Hz, attained the value of 1.29 ± 0.19 (n = 9) and lasted for about 5 min in slices of ‘control’ animals, but was reduced to 1.15 ± 0.12 in ‘trained’ animals (n = 8). Repetitive stimulations delivered at 50 Hz produced LTP with similar extent (1.3 in 50% of the slices) in all three groups. Increasing stimulus frequency to 100 Hz resulted with 60% enhanced LTP in 75% of the slices, but no difference was detected between the three groups. Our data indicate that enhanced odor-learning capability is associated with transient enhancement of synaptic release from the intrinsic fibers in the piriform cortex. Supported by grants from the National Institute for Psychobiology in Israel and the Israeli Ministry for the Arts and Science. ERYTHROCYTE BUT NOT SYNAPTIC HUMAN ACETYLCHOLINESTERASE HYDROLYZES HEROIN TO MORPHINE A. Salmon1, Z. Goren2, Y. Avissar2 and H. Soreq1 1 Dept. of Biological Chemistry, Institute of Life Sciences, Hebrew University of Jerusalem, Jerusalem, Israel; 2 Analytical Laboratory, Division of Identification and Forensic Science, Israel Police In human blood, heroin is rapidly hydrolyzed by sequential deacylation of two ester bonds to yield first 6-monoacetylmorphine (6MAM), then morphine. While serum butyrylcholinesterase (BuChE, EC 3.1.1.8) hydrolyzes heroin to 6-MAM, the bulk of heroin hydrolysis has been reported to take place in the microenvironment of red blood cells, which carry the erythrocyte form of acetylcholinesterase (AChE, EC 3.1.1.7), but not BuChE. Here, we report that in vitro, human erythrocyte AChE hydrolyzes heroin to 6-MAM, with Km of 620 PM, and at the same order of magnitude as that reported for BuChE. Moreover, erythrocyte AChE, but not BuChE, is capable of further hydrolysing 6MAM to morphine, albeit at a considerably slower rate. Both hydrolysis steps are totally blocked by addition of the selective quaternary ammonium AChE inhibitor BW284c51 but not by the BuChE-specific organophosphate inhibitor, iso-OMPA. Intriguingly, solubilized recombinant human AChE of the brain synaptic form, unlike membranebound erythrocyte AChE, was incapable of heroin hydrolysis. These findings reveal a new metabolic role for erythrocyte AChE, the biological function of which is as yet unexplained, and demonstrate distinct properties of the two AChE forms, which previously were catalytically indistinguishable. ION CHANNEL STOCHASTICITY MAY BE A CRITICAL FACTOR IN DETERMINING THE RELIABILITY OF SPIKE TIMING E. Schneidman, B. Freedman and I. Segev Dept. of Neurobiology, Institute of Life Sciences, Institute of Computer Science and Neural Computation Center, Hebrew University of Jerusalem, Jerusalem, Israel The reliability and precision of spike timing in a stochastic Hodgkin-Huxley (H&H) model of an isopotential patch was investigated. The stochasticity was introduced by simulating the activity of each of the single ion channels according to Markovian kinetic models, as suggested by Fitzhugh (J. Cell Comp. Physiol. 66, 111–118 (1965)), and used by Skaugen and Walloe (Acta Physiol. Scand. 107, 343–363 (1979)), Strassberg and DeFelice (Neural Comp. 6, 843–856 (1993)) and others. Distinctly different from the ‘standard’ H&H model, the stochastic model qualitatively imitates the input-dependent reliability characteristics found experimentally for neocortical pyramidal neurons by Mainen and Sejnowski (Science 268, 1503–1508, (1995)). The stochastic model also gives rise to other experimentally-observed phenomena, namely sub-threshold oscillations and ‘missing’ spikes for supra-threshold inputs. Our modeling results, in accordance with the experimental results of Mainen and Sejnowski (1995) and of de Ruyter van Steveninck et al. (Science 275, 1805–1808 (1997)), suggest that the