Shlomo Shapira

Butyrylcholinesterase and acetylcholinesterase prophylaxis against soman poisoning in mice

Human butyrylcholinesterase (BChE, EC 3.1.1.8) or acetylcholinesterase (AChE, EC 3.1.1.7) from fetal bovine serum (FBS), administered i.v. in mice, sequestered at approximately 1:1 stoichiometry the highly toxic anti-ChE organophosphate, 1,2,2-trimethylpropyl methyl-fluorophosphonate (soman). A quantitative linear correlation was demonstrated between blood-ChE levels and the protection conferred by exogeneously administered ChE. Results presented here demonstrate that either human BChE or FBS-AChE is an effective prophylactic measure sufficient to protect mice from multiple LD50S of soman without the administration of post-treatment supportive drugs.

Sarin-induced neuropathology in rats

Sarin, a highly toxic cholinesterase (ChE) inhibitor, administered at near 1 LD50 dose causes severe signs of toxic cholinergic hyperactivity in both the peripheral and central nervous systems (CNS). The present study evaluated acute and long-term neuropathology following exposure to a single LD50 dose of sarin and compared it to lesions caused by equipotent doses of soman described previously. Rats surviving 1 LD50 dose of sarin (95 micrograms/kg; IM), were sacrificed at different time intervals post exposure (4 h-90 days) and their brains were taken for histological and morphometric study. Lesions of varying degrees of severity were found in about 70% of the animals, mainly in the hippocampus, piriform cortex, and thalamus. The damage was exacerbated with time and at three months post exposure, it extended to regions which were not initially affected. Morphometric analysis revealed a significant decline in the area of CA1 and CA3 hippocampal cells as well as in the number of CA1 cells. The neuropathological findings, although generally similar to those described following 1 LD50 soman, differed in some features, unique to each compound, for example, frontal cortex damage was specific to soman poisoning. It is concluded that sarin has a potent acute and long-term central neurotoxicity, which must be considered in the design of therapeutic regimes.

NG-nitro-L-arginine enhances neuronal death following transient forebrain ischemia in gerbils.

Experiments were performed with Mongolian gerbils to study the effect of the specific nitric oxide (NO) synthase inhibitor NG-nitro-L-arginine (L-NNA) on ischemic brain damage induced by 5 min bilateral carotid occlusion. A single i.p. injection of L-NNA did not result in any neuronal loss in the central nervous system. In animals undergoing ischemia, a selective destruction of hippocampal CA1 cells was observed whereas pretreatment with 50 mg/kg L-NNA 4 h before administration of ischemia produced significantly more extensive cell damage in the hippocampus and other brain regions. These findings demonstrate that in this model inhibition of nitric oxide generation augments ischemia-induced neuronal cell injury in the brain.

Aging has a complex effect on a rat model of ischemic stroke.

Stroke in humans is usually associated with advanced age. Nevertheless, almost all animal models of ischemic stroke are based on young animals. The present study was designed to assess the effect of age on the development of ischemic injury in a model of focal brain ischemia in rats. Two age groups of Wistar rats were used: young adult (3 months) and old (24-26 months). Under halothane anesthesia, polyethylene microspheres (50 microm in diameter) were injected into the left common carotid artery following a temporary occlusion of the external carotid artery. Sham-operated rats underwent the same procedure but were injected with an identical volume (100 microl) of saline only. Rats of both experimental groups displayed neurological impairment after surgery. However, contrary to expectation, the young rats were more affected than the old rats. Young rats displayed an abrupt 30% decrement in neurological functions in the first week and then showed a partial functional recovery into a 12% decrement from the second week on. Old rats developed the neurological impairment gradually over a 2-week period (6.3% in the first week and 11% in the second week and thereafter). One month later, rats were tested in a water maze task. Again, performance was more impaired in the young ischemic rats than in the old rats. Histological evaluation revealed more extensive neurological damage in young ischemic as compared to old rats. Thus, although increased age has a critical effect on the evolution of the neurological impairment following focal brain ischemia and stroke, its effects in the rat model were more pronounced in the young animals.

Long-Term Study of Brain Lesions Following Soman, in Comparison to DFP and Metrazol Poisoning

The long-term histopathological effects of acute lethal (95 μg kg-1) and sublethal (56 μg kg-1) doses of soman were studied in rats and were compared to lesions caused by equipotent doses of either another cholinesterase (ChE) inhibitor, DFP (1.8 mg kg-1), or a non-organophosphorus convulsant, metrazol (100 mg kg-1). Severe toxic signs were noted following one LD50 dose administration of all the compounds, yet only soman induced brain lesions. Moreover, even when administered at a sublethal dose (0.5 LD50), soman induced some histological changes without any clinical signs of intoxication. Soman-induced brain lesions were assessed quantitatively using a computerized image analyser. The analysis was carried out for up to 3 months following administration, and a dynamic pattern of pathology was shown. The cortical thickness and area of CA1 and CA3 cells declined significantly as early as 1 week post-exposure. No pathological findings were detected following DFP and metrazol administration. It is therefore suggested that brain lesions are not common for all ChE inhibitors and that convulsions per se are not the only factor leading to brain damage following the administration of soman. The degenerative process (found also with the sublethal dose of soman) might be due to a secondary effect, unrelated to somans clinical toxicity, but leading to long-term brain injuries.

Distribution of 3H-soman in mice

Abstract3H-soman (specific activity 10 Ci/mMol), a potent irreversible cholinesterase inhibitor, was administered IV to mice in a dose of one LD-50, which corresponds to 0.25 mCi/mouse. Animals were sacrificed at 5 min, 2 h and 24 h, and whole body autoradiography was performed. High levels of radioactivity in lung and skin were observed at all time intervals after injection. The central nervous system showed very low concentrations of radioactivity, which remained so for 24 h post-injection. Considerable accumulation of 3H-soman in the urine and gallbladder, and in the intestinal lumen, may indicate these as pathways of soman excretion. Quantitative determinations of radioactivity in various tissue samples were consistent with the above-mentioned findings.It is concluded that the nature of the persistent binding of soman to lung and skin is striking, and may indicate the existence of specific sites for soman depots.

Dose-dependent effect of nitric oxide synthase inhibition following transient forebrain ischemia in gerbils.

The extensive research concerning the interaction between nitric oxide (NO) and ischemic brain tissue has yielded contradictory results. The present study was designed to explore the effect of gradual inhibition of NO production on brain ischemia. Gerbils were administered (i.p.) either saline (control-ischemia), or 5, 10, 25 or 50 mg/kg of NG-nitro-L-arginine (NARG), a specific inhibitor of NO synthase (NOS), and 4 h later were subjected to 5 min of forebrain ischemia. A group receiving 50 mg/kg NARG with sham operation served as a second control (control-NARG) group. Body weights and spontaneous activity were monitored daily until day 6, when the gerbils were sacrificed and their brains processed for histologic-morphometric evaluation. All ischemia groups displayed significant decreases in body weights starting on day 1, as compared to control-NARG (non-ischemic) gerbils. At 24 h post-ischemia spontaneous activity was increased in all ischemia groups in a dose-dependent manner, reaching a peak at 25 mg/kg. Typical ischemia-induced neuronal cell degeneration was observed at the hippocampal CA1 layer in control-ischemia and in each of the dose-groups of 10 mg/kg NARG and above. The 5 mg/kg group displayed damage which was not different from control-NARG, and was milder (P < 0.01) than control-ischemia gerbils and each of the other dose-groups. It is suggested that during ischemia, NO activates a series of processes which are beneficial to brain tissue, whereas an excess amount of NO causes neurotoxic effects.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of fast versus slow rewarming following acute moderate hypothermia in rats

Background: The aim of this study was to compare the biochemical and physiological responses of fast vs. slow rewarming from moderate hypothermia in anaesthetized rats.

Biochemical, pharmacological, and clinical aspects of nitric oxide

Helping rehabilitate Science magazines 1992 Molecule of the Year from its reputation as nothing but an environmental pollutant, the 29 papers give special attention to its characterization in terms of co-factor requirement, substrates, and novel specific inhibitors. They also consider its importanc

Multiparametric Responses to Cortical Spreading Depression Under Nitric Oxide Synthesis Inhibition

Cortical spreading depression (SD) described initially by Leao (1944a) is a multifactorial event affecting the electrical, ionic, metabolic and hemodynamic activities in the brain (Vyskocil et al., 1972; Mayevsky et al., 1974; Bures et al. 1974; Mayevsky and Weiss, 1991). Due to disturbances in the ion homeostasis, the Na+K+-ATPase activity and energy metabolism are stimulated in order to restore the normal extracellular ion levels (Mayevsky et al., 1974; Hansen, 1985; Mayevsky and Weiss, 1991). The hemodynamic response to SD was a challenge to many investigators since the initial observation of dilation of pial vessels (Leao, 1944b). He concluded that the vascular responses are secondary to the local changes in the activity of neural elements. The changes in cerebral blood flow (CBF) just before, during and after the depolarization wave of SD were described by various investigators (Van Harreveld and Stamm, 1952; Van Harreveld and Ochs, 1957; Burevsova, 1957; Hansen et al., 1980; Lauritzen et al., 1982; Mies and Paschen, 1984; Lauritzen, 1984; Lauritzen and Diemer, 1986) and have been reviewed by Lauritzen (1987a,b). In all studies, a large increase in cerebral blood flow was recorded during the wave. Lauritzen and collaborators descried a post-spreading depression wave hypoperfusion, while a preceding vasoconstriction (immediately before the wave) was not established or proved. The mechanism behind the changes in CBF due to the SD wave is not clear although recently nitric oxide NO was proposed to be involved (Goadsby et al., 1992; Duckrow 1993). NO was suggested as an important factor in CBF regulation (Beckman et al., 1991; Iadecola et al., 1994; Irikura et al.,1994) as well as having direct effects on neuronal elements (Culotta and Koshland 1992; Mayer et al., 1992). In order to