Neuropathology of organophosphate-induced delayed neuropathy (OPIDN) in young chicks

To examine the phenomenon of apparent age resistance of young chicks to organophosphate-induced delayed neuropathy (OPIDN), groups of either 2- or 10-week-old chicks were exposed subcutaneously daily for 4 days to the neuropathic organophosphate (OP), di-isopropylfluorophosphate (DFP, 1 mg/kg), the non-neuropathic OP, paraoxon (PO, 0.25 mg/kg) or atropine (20 mg/kg). Subsequently, all birds were examined at post-exposure intervals (calculated from the last day of exposure) for up to 56 days for neurological deficits and morphological lesions in the central and peripheral nervous systems (CNS, PNS). Clinically, none of the birds in the 2-week-old groups, or in the 10-week-old PO or atropine exposed groups had neurological deficits. However, all birds in the 10-week-old DFP exposed group developed ataxia by 7 days post-exposure (DPE) and then progressive paralysis. Therefore, all birds in the 10-week-old groups were killed at 14 DPE. Pathologically, the 2-week-old DFP exposed chicks had increasingly severe lesions of Wallerian-like degeneration predominantly in the spinal cord from 7 DPE and subsequently. In the 10-week-old DFP exposed chicks, the degenerative lesions of OPIDN were first detected in the CNS at 3 DPE and then with equally increasing severity in the CNS and PNS up to 14 DPE. A higher incidence of neuronal necrosis and chromatolysis in ventral motor horn neurons of spinal cord grey matter and in dorsal root ganglia occurred in both the DFP exposed age groups compared with those lesions in other groups. These results demonstrate that after neuropathic DFP exposure, 2-week-old chicks develop pathological lesions in the spinal cord without neurological deficits. In both age groups, onset of degenerative lesions in the spinal cord preceeded those in the PNS. The claim of apparent age resistance of chicks to OPIDN needs to be re-evaluated.     

Age sensitivity to organophosphate-induced delayed polyneuropathy. Biochemical and toxicological studies in developing chicks

Young animals are resistant to organophosphate-induced delayed polyneuropathy (OPIDP). The putative target protein in the nervous system for initiation of OPIDP in the adult hen is an enzyme called Neuropathy Target Esterase (NTE), which is dissected by selective inhibitors among nervous tissue esterases hydrolysing phenyl valerate (PV). We report here that the pool of PV-esterases sensitive to paraoxon was different in peripheral nerves of chicks as compared to that of hens while that of brain and spinal cord was not. NTE activity decreased with age in brain, spinal cord and peripheral nerve, but its sensitivity to several inhibitors remained unchanged. In the adult hen more than 70% inhibition of peripheral nerve NTE by neuropathic OPs is followed by deficit of retrograde axonal transport, axonal degeneration and paralysis. Similar NTE inhibition in 40-day-old or younger chicks however is not followed by changes in retrograde axonal transport nor by OPIDP. Chicks aged 60 to 80 days are only marginally sensitive to a single dose of DFP otherwise clearly neuropathic to hens. In vitro and in vivo phosphorylation by DFP and subsequent aging of brain NTE is similar both in chicks and in hens. The recovery of NTE activity monitored in vivo after inhibition by DFP is faster (half-life of about 3 days) in chick peripheral nerves as compared to chick brain, hen brain and hen peripheral nerve (half-life of about 5 days). It is concluded that the reduced sensitivity to OPIDP in chicks is not due to differences in OP-NTE interactions. The resistance might be explained by a more efficient repair mechanism, as suggested by the faster recovery of peripheral nerve NTE activity.     

Inhibition of lymphocytic neuropathy target esterase predicts the development of organophosphate-induced delayed polyneuropathy

Neuropathy Target Esterase (NTE) is the molecular target in the nervous system for organophosphorus esters (OP) when they cause delayed polyneuropathy. Some NTE activity was recently found also in blood lymphocytes. An unsuccessful suicide attempt with the widely used pesticide chlorpyrifos (0,0-diethyl-0-3,5,6,-trichloro-2-pyridyl phosphorothioate) is reported, where prior inhibition of lymphocytic NTE correlates with the delayed development of polyneuropathy. A 42-year-old man drank approximately 300 mg/kg chlorpyrifos. The subsequent severe cholinergic syndrome lasted for 17 days with varying degrees of severity. Thirty days after intoxication the clinical and electrophysiological examination of the peripheral nervous system was normal but lymphocytic NTE was about 60% inhibited. On day 43 the patient began to complain of paresthesia and leg weakness. Clinical examination, electrophysiology and a nerve biopsy revealed signs of a peripheral polyneuropathy, axonal in type. This case report indicates that measurement of lymphocytic NTE might be used as a clinical test to predict the development of OP-induced delayed polyneuropathy.     

Clinical expression of organophosphate-induced delayed polyneuropathy in rats.

Single doses of certain organophosphates (OP), such as dibutyl-2,2-dichlorovinyl phosphate (DBDCVP) cause organophosphate-induced delayed polyneuropathy (OPIDP) in hens. Clinical effects correlate with inhibition of neuropathy target esterase (NTE) which is considered the target for this toxicity. Pre-treatment with non-neuropathic NTE inhibitors, such as phenylmethanesulfonyl fluoride (PMSF), protects from OPIDP. However, when given after OPs, these compounds promote OPIDP. Chicks are relatively resistant to OPIDP despite high NTE inhibition. It has also always been reported that rats represent a species which is resistant to OPIDP and that they might develop morphological but not clinical signs of OPIDP. We report here that clinical OPIDP can be produced in 3.5- and 6-month-old rats by DBDCVP (5 mg/kg s.c.) and that it correlates with high (> 90%) NTE inhibition. When PMSF (120 mg/kg s.c. x 2) was given after DBDCVP, OPIDP was promoted. Pretreatment with PMSF protected from OPIDP. We conclude that resistance to OPIDP in the rat is age-related, as it is in the hen.     

Characterization of delayed neurotoxicity in the mouse following chronic oral administration of tri-o-cresyl phosphate

The sensitivity of the mouse to organophosphorus-induced delayed neurotoxicity (OPIDN) has been investigated. One group of five mice received two single 1000-mg/kg po doses of tri-o-cresyl phosphate (TOCP) at a 21-day interval (on Days 1 and 21 of the study); a second group of five mice was given 225 mg/kg of TOCP daily for 270 days. A third group of five animals served as an untreated control. All animals were killed 270 days after the start of the experiment. Daily po dosing of 225 mg/kg TOCP caused a decrease in body weight gain, muscle wasting, weakness, and ataxia which progressed to severe hindlimb paralysis at termination. On the other hand, po administration of two single 1000-mg/kg doses of TOCP at a 21-day interval produced no observable adverse effects. Brain acetylcholinesterase (AChE) and neurotoxic esterase (NTE) activity were 35 and 10% of the control, respectively, in daily dosed animals while AChE and NTE in mice receiving two single 1000-mg/kg doses of TOCP were not significantly altered from the control group. Plasma butyrylcholinesterase activity was 12% of the control group in daily dosed animals. Hepatic microsomal enzyme activities of aniline hydroxylase and p-chloro-N-methylaniline demethylase and NADPH-cytochrome P-450 content in daily dosed animals were increased (141 to 161% of the control group) when compared to controls and mice receiving two single 1000-mg/kg doses of TOCP; the latter being not significantly different from each other. Degeneration of the axon and myelin of the spinal cord and sciatic fascicle were observed and were consistent with OPIDN. This study demonstrates that chronic dosing of TOCP produces OPIDN and induces hepatic microsomal enzyme activity in mice. It is concluded that while the mouse is susceptible to OPIDN, it is a less sensitive and a less appropriate test animal for studying this effect when compared to the adult hen.     

Effect of route of administration on drug disposition

While the first-pass effect is a fairly generally understood phenomenon in the gross sense, it is not commonly appreciated that the rate at which the drug is delivered to the liver can influence the fraction of the administered dose which reaches the fluids of distribution. The drug molecules being absorbed via the hepatic portal vein intermix with previously absorbed drug recycling in the hepatic artery. The concentration of the drug returning via the hepatic artery is affected by the volume of distribution of the drug into body tissues (and the relative partition of the drug between the blood and the tissues). If one presumes that liver uptake is controlled by a Michaelis-Menten process of metabolism, it can be readily shown that the extent of extraction of the drug by the liver is controlled not only by the Michaelis-Menten constants, but also by the concentration of the drug transitting the liver per unit time. The effects of dose, rate of absorption, partitioning, and tissue distribution characteristics are systematically discussed and referred to in relation to selected examples of drugs in common use.The work reported here was supported in part by grant NIGMS16496-05.Presented by S. Reigelman at the Conference on Pharmacology and Pharmacokinetics: Problems and Perspectives, October 30–November 1, 1972, at the Fogarty International Center, National Institutes of Health, Bethesda, Maryland. This paper, in a slightly different format, will be published in the Proceedings of the Conference by Plenum Press, New York.     

Systemic administration of 4-chlorokynurenine prevents quinolinate neurotoxicity in the rat hippocampus

The synthetic compound 4-chlorokynurenine has been shown to be enzymatically transaminated to the selective glycine_B receptor antagonist 7-chlorokynurenate. Since 4-chlorokynurenine, in contrast to 7-chlorokynurenate, readily penetrates the blood–brain barrier, the present study evaluated its neuroprotective properties after systemic administration in rats. Intrahippocampal injection of the NMDA receptor agonist quinolinate (15 nmol/1 μl) was used as the neurotoxic paradigm. Serum and hippocampal tissue measurements confirmed that 4-chlorokynurenine serves as an effective pro-drug of 7-chlorokynurenate both in the periphery and in the brain. These studies and complementary hippocampal microdialysis experiments compared the effects of single and repeated injections of 4-chlorokynurenine (50 or 200 mg/kg, intraperitoneal (i.p.), 10 min prior to an intrahippocampal quinolinate injection; or 50 mg/kg, i.p., 10 min before and 30, 120 and 360 min after quinolinate). With the multiple-dosing regimen, extracellular 7-chlorokynurenate levels in the hippocampus reached a maximum of approximately 750 nM 7 h after quinolinate and gradually decreased with a half-life of about 3 h. In contrast, a single injection of 200 mg/kg 4-chlorokynurenine resulted in a considerably shorter rise in extracellular 7-chlorokynurenate without yielding higher peak levels. In separate animals, repeated treatment with 50 mg/kg 4-chlorokynurenine, but not a single injection of 200 mg/kg of the pro-drug, provided total protection against quinolinate-induced excitotoxicity. These data suggest that a prolonged and functionally relevant blockade of hippocampal glycine_B receptors can be achieved after the systemic administration of 4-chlorokynurenine.     

Variation of neurotoxicity of L- and D-2,4-diaminobutyric acid with route of administration

When typical neurotoxicity symptoms (hyperirritability, tremors and convulsions) were produced by ip or intracisternal injection of l- or d-2,4-diaminobutyric acid to rats, the concentration of diaminobutyric acid in brain always exceeded 1 mm. The occurrence of toxicity was not dependent on a change in the concentration of any normal brain amino acid. The structurally related 1,3-diaminopropane and 1,4-diaminobutane are potent neurotoxins.     

Neuregulin-1 is neuroprotective in a rat model of organophosphate-induced delayed neuronal injury

Current medical countermeasures against organophosphate (OP) nerve agents are effective in reducing mortality, but do not sufficiently protect the CNS from delayed brain damage and persistent neurological symptoms. In this study, we examined the efficacy of neuregulin-1 (NRG-1) in protecting against delayed neuronal cell death following acute intoxication with the OP diisopropylflurophosphate (DFP). Adult male Sprague-Dawley rats were pretreated with pyridostigmine (0.1 mg/kg BW, i.m.) and atropine methylnitrate (20 mg/kg BW, i.m.) prior to DFP (9 mg/kg BW, i.p.) intoxication to increase survival and reduce peripheral signs of cholinergic toxicity but not prevent DFP-induced seizures or delayed neuronal injury. Pretreatment with NRG-1 did not protect against seizures in rats exposed to DFP. However, neuronal injury was significantly reduced in most brain regions by pretreatment with NRG-1 isoforms NRG-EGF (3.2 μg/kg BW, i.a) or NRG-GGF2 (48 μg/kg BW, i.a.) as determined by FluroJade-B labeling in multiple brain regions at 24 h post-DFP injection. NRG-1 also blocked apoptosis and oxidative stress-mediated protein damage in the brains of DFP-intoxicated rats. Administration of NRG-1 at 1 h after DFP injection similarly provided significant neuroprotection against delayed neuronal injury. These findings identify NRG-1 as a promising adjuvant therapy to current medical countermeasures for enhancing neuroprotection against acute OP intoxication.     

The effect of testosterone or estradiol on the development of TOCP-induced delayed neurotoxicity in immature broiler-breed cockerels

Immature cockerels were susceptible to OPIDN when dosed with TOCP. Using 30 broiler-breed cockerels, 6 w old, 10 birds each received 28 daily im injections of either 50 mg estradiol, 100 mg testosterone or 0.1 ml of vehicle. At 7 w of age, 5 birds in each of the 3 groups received a single oral dose of 500 mg TOCP/kg body weight, while the remaining 5 birds/groups were given corn oil. The birds were observed daily for 14 d beginning on day 8 post-TOCP exposure for the development of clinical signs characteristic of OPIDN. At 21 d post-TOCP, all birds were killed and the adrenal gland and testes were prepared for histopathology of the birds that received TOCP, 1 of 5 that were given testosterone and 2 of 5 that received estradiol had signs typical of OPIDN. All of the 5 birds that received TOCP alone showed OPIDN signs. The testes of the TOCP-exposed birds that showed clinical signs had reductions in the size of the seminiferous tubules and no evidence of spermatogenic activity. This study demonstrated that sex hormones can modulate the clinical effects of TOCP in immature cockerels through unknown mechanisms that are similar to those reported for corticosterone in adult chickens.     

The effect of hepatic mixed-function oxidase enzyme inducers on the development of tri-o-tolyl phosphate-induced delayed neurotoxicity

Adult White Leghorn hens were divided into 15 groups of 10 birds each. Five groups received the prototype mixed-function oxidase (MFO) enzyme inducer phenobarbital (PB) at a dosage of 50 mg/kg body weight for 3 consecutive days by intraperitoneal (i.p.) injection, 5 groups received the MFO enzyme inducer beta-naphthoflavone (beta NF) at 20 mg/kg body weight for 2 consecutive days by i.p. injection, while the remaining 5 groups did not receive an inducer. At 24 h after the last injection, the birds received a single oral dose of tri-o-tolyl phosphate (TOTP) (an organophosphate that produces delayed neurotoxicity after metabolic activation by the MFO system) in doses of 62.5, 125, 250, or 500 mg/kg body weight. Corn oil served as the vehicle control. At 48 h after the administration of TOTP, half the birds in each of the 15 groups were killed for determination of whole-brain neurotoxic esterase (NTE) activity. The remaining birds were observed for the subsequent 19 d for onset of clinical signs characteristic of delayed neurotoxicity. Birds receiving beta NF prior to TOTP were protected by the inducer when compared to birds receiving PB + TOTP or TOTP alone. This was indicated by less severe clinical signs as well as less inhibition of whole-brain NTE activity. The protective effect offered by beta NF may be due to induction of enzymes responsible for the inactivation of the neurotoxic metabolite.     

Effect of route of administration and distribution on drug action

The extent and time course of drug action can be markedly affected by the route of drug administration into the patient as well as the pattern of drug distribution within the patient. Drugs which are rapidly cleared by hepatic processes will show a decreased extent of availability following oral administration due to metabolism of drug on its first pass through the liver. The magnitude of this first pass will depend on the blood flow to the liver and the intrinsic clearing ability of the liver (i.e., the ability of the organ to eliminate the drug independent of the rate at which drug is brought to the organ). Drug distribution in the patient will depend on the blood flow to various sites in the body as well as the partition coefficient of the drug between the blood and the distributive organs. Protein binding both in the plasma and in the tissues will markedly affect this distribution. However, free drug concentrations are generally believed to be the effective determinant in drug therapy. Often a redistribution due to changes in protein binding will have little effect on the therapeutic efficacy since, although total drug distribution changes, free concentrations in the plasma remain essentially similar.This article is extracted from the forthcoming bookModern Pharmaceutics: Drug Product Design and Evaluation, edited by Gilbert S. Banker and Christopher T. Rhodes, copyright 1978, Marcel Dekker, Inc. Published with permission.     

Effect of delayed administration of activated charcoal on nortriptyline absorption

Summary Activated charcoal is known to reduce the absorption of therapeutic doses of nortriptyline in vivo when administered 30 min after drug ingestion. In a group of volunteers, one sachet (10 g) of a new activated charcoal preparation, Medicoal was found to produce a highly significant reduction in nortriptyline absorption when given as long as four hours after nortriptyline dosing. Activated charchoal may therefore be useful in the treatment of tricyclic antide-pressant poisoning even if a delay of several hours ensues before medical help is sought.     

康复训练联合神经肌肉电刺激对有机磷迟发性神经病的疗效

目的 观察康复训练联合神经肌肉电刺激(NMES)对有机磷迟发性神经病(OPIDP)的疗效.方法 44例OPIDP患者,随机分为对照组和试验组,每组22例.对照组入院后开始康复训练;试验组患者在此基础上增加NMES治疗.比较两组患者简式Fugl-Meyer评定量表(FMA)评分、改良Barthel指数(MBI)评分、Be...     

Protective effects of IRFI-042 in monensin induced neurotoxicity in chicks

Monensin, a well known ionophore antibiotic, may cause severe damage in neuronal cells by altering Na+/K+-ATPase and Ca2+-ATPase. We investigated whether IRFI-042, a synthetic analogue of vitamin E, may block lipid peroxidation in neuronal cells and protect against monensin neurotoxicity in chicks. Monensin toxicity was induced in chicks by once-daily administration (150 mg/kg by oral gavages), for 8 days. Sham animals received a saline solution and were used as controls. All animals were randomized to receive either IRFI-042 (20 mg/kg) or its vehicle. Survival rate, brain lipid peroxidation, mRNA for neuronal and inducible nitric oxide synthases (nNOS and iNOS) and brain histological evaluations, including immunohistochemical expression of nNOS and iNOS were performed. Monensin administration decreased survival rate, induced behavioural changes, increased brain lipid peroxidation, reduced brain nNOS mRNA and immunostaining and enhanced iNOS mRNA and immunostaining in the brain in chicks. IRFI-042 significantly improved the survival rate and counteracted monensin-induced changes in chick brains. Our data suggest that monensin is responsible of neurotoxicity in chicks by inducing oxidative stress/lipid peroxidation and that IRFI-042 might represent a useful pharmacological approach to protect against the neuronal damage induced by this monovalent carboxylic ionophorous polyether antibiotic.     

The effects of drug metabolism inducers on the delayed neurotoxicity and disposition of tri-o-cresyl phosphate in hens following a single intravenous administration

Intravenous (iv) administration of tri-o-cresyl phosphate (TOCP) caused a delayed neurotoxic effect in hens similar to that produced by oral and dermal administration. The iv ED50s for producing ataxia and paralysis were estimated to be 15.9 mg/kg and 31.7 mg/kg respectively. The tissue disposition of unaltered TOCP was determined in hens following a single iv injection of 40 mg/kg of TOCP. One hour (hr) after the injection, the leg muscle contained the highest concentration, 26.99 micrograms/g fresh weight followed by the adipose tissue. Among the nerve tissues, the sciatic nerve had the highest concentration, 9.63 micrograms/g followed by the spinal cord and the brain. Except the adipose tissue and the sciatic nerve, the concentration in all analyzed tissues dropped below 1.0 microgram/g (ml) after 24 hr. An unidentified metabolite appeared in bile taken 1 and 3 hr after the injection. Pretreatment of hens with 3-methylcolathrene (3-MC) and beta-naphthoflavone (B-NF) protected against the TOCP-induced delayed neurotoxicity, whereas phenobarbital (PB) failed to protect against the neurotoxicity. Plasma creatine phosphokinase (CK) activity in paralytic birds increased approximately 4 times of the control or symptomless hens on the 21st day. 3-MC-, B-NF- and PB-treatment depressed substantially the concentration of unaltered TOCP in brain and plasma 1 hr after iv dosing with 40 mg/kg of TOCP. Only B-NF pretreatment lowered the level of TOCP in spinal cord. There was no effect of these inducers on the level of TOCP in sciatic nerve and adipose tissue. B-NF and 3-MC lowered significantly the TOCP level in leg muscle, whereas PB had no such effect. More attention should be paid to the role of TOCP in muscle, especially to the leg muscle, judging from the present toxicological and metabolic studies.