A method for quantitative risk appraisal for pesticide risk assessments.

Pesticide risk assessments are fraught with uncertainties that arise from the process of estimating exposure to and toxicity of chemicals. Regulatory agencies resolve those uncertainties in a health-protective (conservative) manner, typically acknowledging only inter- and intraspecies uncertainties quantitatively. Other uncertainties may be acknowledged qualitatively, but those safety factors (SF) are not enumerated. Quantitative risk appraisal may be used to enumerate the multiplicative SF generated by conservative assumptions regarding uncertainties. The magnitude of SF derived from decision points dealing with historically unquantified uncertainty may far exceed explicit SF used to gauge acceptable margins of exposure (MoE). Examination of the basis for some previously unenumerated SF may justify potential changes in regulatory practices and policies. Using past risk assessments of 3 pesticides (mevinphos, parathion, and methyl iodide) for which the California Department of Pesticide Regulation found unacceptable risk as examples, the previously unquantified SF ranged from 47 to 1 × 106 for scenarios involving handlers, reentry workers, and bystanders.

Pro-life role for c-Jun N-terminal kinase and p38 mitogen-activated protein kinase at rostral ventrolateral medulla in experimental brain stem death.

BACKGROUND: Based on an experimental brain stem death model, we demonstrated previously that activation of the mitogen-activated protein kinase kinase 1/2 (MEK1/2)/extracellular signal-regulated kinase 1/2 (ERK1/2)/ mitogen-activated protein kinase signal-interacting kinase 1/2 (MNK1/2) cascade plays a pro-life role in the rostral ventrolateral medulla (RVLM), the origin of a life-and-death signal detected from systemic arterial pressure, which sequentially increases (pro-life) and decreases (pro-death) to reflect progressive dysfunction of central cardiovascular regulation during the advancement towards brain stem death in critically ill patients. The present study assessed the hypothesis that, in addition to ERK1/2, c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38MAPK), the other two mammalian members of MAPKs that are originally identified as stress-activated protein kinases, are activated specifically by MAPK kinase 4 (MAP2K4) or MAP2K6 and play a pro-life role in RVLM during experimental brain stem death. We further delineated the participation of phosphorylating activating transcriptional factor-2 (ATF-2) and c-Jun, the classical transcription factor activated by JNK or p38MAPK, in this process. RESULTS: An experimental model of brain stem death that employed microinjection of the organophosphate insecticide mevinphos (Mev; 10 nmol) bilaterally into RVLM of Sprague-Dawley rats was used, alongside cardiovascular, pharmacological and biochemical evaluations. Results from ELISA showed that whereas the total JNK, p38MAPK, MAP2K4 and MAP2K6 were not affected, augmented phosphorylation of JNK at Thr183 and Tyr185 and p38MAPK at Thr180 and Tyr182, accompanied by phosphorylation of their upstream activators MAP2K4 at Ser257 and Thr261 and MAP2K6 at Ser207 and Thr211 in RVLM occurred preferentially during the pro-life phase of experimental brain stem death. Moreover, the activity of transcription factors ATF-2 at Thr71 and c-Jun at Ser73, rather than Elk-1 at Ser383 in RVLM were also augmented during the pro-life phase. Furthermore, pretreatment by microinjection into the bilateral RVLM of specific JNK inhibitors, JNK inhibitor I (100 pmol) or SP600125 (5 pmol), or specific p38MAPK inhibitors, p38MAPK inhibitor III (500 pmol) or SB203580 (2 nmol), exacerbated the depressor effect and blunted the augmented life-and-death signal exhibited during the pro-life phase. On the other hand, pretreatment with the negative control for JNK or p38MAPK inhibitor, JNK inhibitor I negative control (100 pmol) or SB202474 (2 nmol), was ineffective in the vehicle-controls and Mev-treatment groups. CONCLUSIONS: Our results demonstrated that activation of JNK or p38MAPK in RVLM by their upstream activators MAP2K4 or MAP2K6 plays a preferential pro-life role by sustaining the central cardiovascular regulatory machinery during experimental brain stem death via phosphorylation and activation of nuclear transcription factor ATF-2 or c-Jun.

Alternative bioassay for the detection of saxitoxin using the speckled cockroach (Nauphoeta cinerea).

Paralytic shellfish poisoning (PSP) toxins produced by cyanobacteria pose a risk to public health as they occur in drinking water reservoirs and recreational lakes and accumulate in the food chain. One of these PSP toxins, saxitoxin (STX) is one of the most toxic nonprotein substances known. Accordingly, there is a requirement to monitor for these toxins. The standard bioassay used to detect these toxins is the mouse bioassay; however, its use is constrained by animal ethics guidelines and practical considerations. Reported here is the use of the globally distributed speckled cockroach Nauphoeta cinerea as a bioassay test organism for the selective detection of PSP toxicity of Anabaena circinalis aqueous extract and STX. N. cinerea was shown to be tolerant to pure cylindrospermopsin (CYN) and microcystin-LR (MC-LR) at doses 10-fold greater than mouse LD50 values while being sensitive to STX. Similarly, N. cinerea was shown to be tolerant of toxin-containing aqueous extracts of Cylindrospermopsis raciborskii, Microcystis aeruginosa, and Nodularia spumigena while being sensitive to A. circinalis. Peak sensitivity to STX was 60 min postinjection with a KD50 of 31.2 ng/g body weight. While this was approximately 3-fold less sensitive than the mouse bioassay, the insect test organism was around 34-fold smaller in mass than a mouse (20 g); thus one-tenth the amount of toxin in absolute quantity was required to reach an ED50 level. The N. cinerea bioassay presents a selective test for PSP toxicity that is rapid, economical, efficient, and simple to perform.

Outcomes of elderly patients with organophosphate intoxication.

This study analysed the clinical patterns and outcomes of elderly patients with organophosphate intoxication. A total of 71 elderly patients with organophosphate poisoning were seen between 2008 and 2017. Patients were stratified into two subgroups: survivors (n = 57) or nonsurvivors (n = 14). Chlorpyrifos accounted for 33.8% of the cases, followed by methamidophos (12.7%) and mevinphos (11.3%). Mood, adjustment and psychotic disorder were noted in 39.4%, 33.8% and 2.8% of patients, respectively. All patients were treated with atropine and pralidoxime therapies. Acute cholinergic crisis developed in all cases (100.0%). The complications included respiratory failure (52.1%), aspiration pneumonia (50.7%), acute kidney injury (43.7%), severe consciousness disturbance (25.4%), shock (14.1%) and seizures (4.2%). Some patients also developed intermediate syndrome (15.5%) and delayed neuropathy (4.2%). The nonsurvivors suffered higher rates of hypotension (P < 0.001), shock (P < 0.001) and kidney injury (P = 0.001) than survivors did. Kaplan-Meier analysis indicated that patients with shock suffered lower cumulative survival than did patients without shock (log-rank test, P < 0.001). In a multivariate-Cox-regression model, shock was a significant predictor of mortality after intoxication (odds ratio 18.182, 95% confidence interval 2.045-166.667, P = 0.009). The mortality rate was 19.7%. Acute cholinergic crisis, intermediate syndrome, and delayed neuropathy developed in 100.0%, 15.5%, and 4.2% of patients, respectively.

Heme oxygenase-1 plays a pro-life role in experimental brain stem death via nitric oxide synthase I/protein kinase G signaling at rostral ventrolateral medulla.

BACKGROUND: Despite its clinical importance, a dearth of information exists on the cellular and molecular mechanisms that underpin brain stem death. A suitable neural substrate for mechanistic delineation on brain stem death resides in the rostral ventrolateral medulla (RVLM) because it is the origin of a life-and-death signal that sequentially increases (pro-life) and decreases (pro-death) to reflect the advancing central cardiovascular regulatory dysfunction during the progression towards brain stem death in critically ill patients. The present study evaluated the hypothesis that heme oxygnase-1 (HO-1) may play a pro-life role as an interposing signal between hypoxia-inducible factor-1 (HIF-1) and nitric oxide synthase I (NOS I)/protein kinase G (PKG) cascade in RVLM, which sustains central cardiovascular regulatory functions during brain stem death. METHODS: We performed cardiovascular, pharmacological, biochemical and confocal microscopy experiments in conjunction with an experimental model of brain stem death that employed microinjection of the organophosphate insecticide mevinphos (Mev; 10 nmol) bilaterally into RVLM of adult male Sprague-Dawley rats. RESULTS: Western blot analysis coupled with laser scanning confocal microscopy revealed that augmented HO-1 expression that was confined to the cytoplasm of RVLM neurons occurred preferentially during the pro-life phase of experimental brain stem death and was antagonized by immunoneutralization of HIF-1α or HIF-1ß in RVLM. On the other hand, the cytoplasmic presence of HO-2 in RVLM neurons manifested insignificant changes during both phases. Furthermore, immunoneutralization of HO-1 or knockdown of ho-1 gene in RVLM blunted the augmented life-and-death signals exhibited during the pro-life phase. Those pretreatments also blocked the upregulated pro-life NOS I/PKG signaling without affecting the pro-death NOS II/peroxynitrite cascade in RVLM. CONCLUSIONS: We conclude that transcriptional upregulation of HO-1 on activation by HIF-1 in RVLM plays a preferential pro-life role by sustaining central cardiovascular regulatory functions during brain stem death via upregulation of NOS I/PKG signaling pathway. Our results further showed that the pro-dead NOS II/peroxynitrite cascade in RVLM is not included in this repertoire of cellular events.

Redox-active DJ-1 sustains brainstem cardiovascular regulation via maintenance of mitochondrial function during mevinphos intoxication.

DJ-1 (also known as PARK7) is a redox-active protein that protects against oxidative stress. This study evaluated the hypothesis that DJ-1 sustains brainstem cardiovascular regulation via maintaining mitochondrial function in the rostral ventrolateral medulla (RVLM), a brainstem site known to maintain blood pressure and sympathetic vasomotor tone, during cardiovascular depression elicited by the organophosphate insecticide mevinphos. In Sprague-Dawley rats, intravenous administration of mevinphos (640 µg kg-1) resulted in progressive hypotension, accompanied by an increase (Phase I) followed by a decrease (Phase II) of an experimental index for spontaneous baroreflex-mediated sympathetic vasomotor tone, alongside elevation in mitochondrial superoxide levels in the RVLM. There was concurrent activation of DJ-1 induced by oxidative stress in the RVLM, which was causally and temporally related to translocation of DJ-1 to mitochondria, reduction in mitochondrial membrane potential, increase in cytosolic apoptosis-inducing factor level, and apoptotic cell death in this brainstem site. Loss-of-function by immunoneutralization of DJ-1 in the RVLM significantly exacerbated those biochemical and cellular events, enhanced the progressive hypotension, diminished the increased and augmented the decreased spontaneous baroreflex-mediated sympathetic vasomotor tone respectively during Phases I and II, and heightened lethality during mevinphos intoxication. We conclude that DJ-1 in the RVLM sustains brainstem cardiovascular regulation induced by mevinphos via maintaining mitochondrial function.

Muscarinic receptor-independent activation of cyclic adenosine monophosphate-dependent protein kinase in rostral ventrolateral medulla underlies the sympathoexcitatory phase of cardiovascular responses during mevinphos intoxication in the rat.

As inhibitors of acetylcholinesterase, clinical presentations of poisoning from organophosphate compounds are generally believed to entail overstimulation by the accumulated acetylcholine on muscarinic receptors at peripheral and central synapses. That some patients still yielded to acute organophosphate poisoning despite repeated dosing of atropine suggests that cellular mechanisms that are independent of muscarinic receptor activation may also be engaged in organophosphate poisoning. The present study was undertaken to test the hypothesis that muscarinic receptor-independent activation of cyclic adenosine monophosphate-dependent protein kinase A (PKA) in rostral ventrolateral medulla (RVLM), a medullary site where sympathetic vasomotor tone originates and where the organophosphate poison mevinphos (Mev) acts, is involved in the cardiovascular responses exhibited during organophosphate intoxication. In Sprague-Dawley rats, microinjection bilaterally of Mev (10 nmol) into the RVLM significantly augmented PKA activity in ventrolateral medulla that was not antagonized by coadministration of an equimolar concentration (1 nmol) of atropine or selective muscarinic receptor type M1 (pirenzepine), M2 (methoctramine), M3 (4-diphenyl-acetoxy-N-dimethylpiperidinium), or M4 (tropicamide) inhibitor. Comicroinjection of two selective PKA antagonists (100 pmol), N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide and (9R,10S,12S)-2,3,9,10,11,12-hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolol[1,2,3-fg:3',2',1'-kl]pyrrolo[3,4-1][1,6]benzodiazocine-10-carboxylic acid, significantly blunted the initial sympathoexcitatory cardiovascular response and the accompanying augmentation of nitric oxide synthase (NOS I) expression in the ventrolateral medulla exhibited during Mev intoxication; the secondary sympathoinhibitory phase and associated elevation in NOS II expression were unaffected. We conclude that whereas a muscarinic receptor-independent augmentation of PKA activity in the ventrolateral medulla was manifested throughout acute Mev intoxication, this activation was preferentially involved in the sympathoexcitatory phase by an upregulation of NOS I expression.

PTEN, a negative regulator of PI3K/Akt signaling, sustains brain stem cardiovascular regulation during mevinphos intoxication.

Activation of PI3K/Akt signaling, leading to upregulation of nitric oxide synthase II (NOS II)/peroxynitrite cascade in the rostral ventrolateral medulla (RVLM), the brain stem site that maintains blood pressure and sympathetic vasomotor tone, underpins cardiovascular depression induced by the organophosphate pesticide mevinphos. By exhibiting dual-specificity protein- and lipid-phosphatase activity, phosphatase and tensin homolog (PTEN) directly antagonizes the PI3K/Akt signaling by dephosphorylation of phosphatidylinositol-3,4,5-trisphosphate, the lipid product of PI3K. Based on the guiding hypothesis that PTEN may sustain brain stem cardiovascular regulation during mevinphos intoxication as a negative regulator of PI3K/Akt signaling in the RVLM, we aimed in this study to clarify the mechanistic role of PTEN in mevinphos-induced circulatory depression. Microinjection bilaterally of mevinphos (10 nmol) into the RVLM of anesthetized Sprague-Dawley rats induced a progressive hypotension and a decrease in baroreflex-mediated sympathetic vasomotor tone. There was progressive augmentation in PTEN activity as reflected by a decrease in the oxidized form of PTEN in the RVLM during mevinhpos intoxication, without significant changes in the mRNA or protein level of PTEN. Loss-of-function manipulations of PTEN in the RVLM by immunoneutralization, pharmacological blockade or siRNA pretreatment significantly potentiated the increase in Akt activity or NOS II/peroxynitrite cascade in the RVLM, enhanced the elicited hypotension and exacerbated the already reduced baroreflex-mediated sympathetic vasomotor tone. We conclude that augmented PTEN activity via a decrease of its oxidized form in the RVLM sustains brain stem cardiovascular regulation during mevinphos intoxication via downregulation of the NOS II/peroxynitrite cascade as a negative regulator of PI3K/Akt signaling.

Cholinergic-receptor-independent dysfunction of mitochondrial respiratory chain enzymes, reduced mitochondrial transmembrane potential and ATP depletion underlie necrotic cell death induced by the organophosphate poison mevinphos.

Our current understanding of the nature of cell death that is associated with fatal organophosphate poisoning and the underlying cellular mechanisms is surprisingly limited. Taking advantage of the absence in an in vitro system of acetylcholinesterase, the pharmacological target of organophosphate compounds, the present study evaluated the hypothesis that the repertoire of cholinergic receptor-independent cellular events that underlie fatal organophosphate poisoning entails induction of mitochondrial dysfunction, followed by bioenergetic failure that leads to necrotic cell death because of ATP depletion. Pheochromocytoma PC12 cells incubated with the organophosphate pesticide mevinphos (0.4 or 4mumol) for 1 or 3h underwent a dose-related and time-dependent loss of cell viability that was not reversed by muscarinic (atropine) or nicotinic (mecamylamine) blockade. This was accompanied by depressed NADH cytochrome c reductase, succinate cytochrome c reductase or cytochrome c oxidase activity in the mitochondrial respiratory chain, reduced mitochondrial transmembrane potential, decreased ATP concentration, elevated ADP/ATP ratio, increased lactate dehydrogenase release and necrotic cell death. We conclude that Mev induces cholinergic receptor-independent necrotic cell death by depressing the activity of Complexes I to IV in the mitochondrial respiratory chain, eliciting reduction in mitochondrial transmembrane potential, depleting intracellular ATP contents and damaging cell membrane integrity.

Neuroprotective role of coenzyme Q10 against dysfunction of mitochondrial respiratory chain at rostral ventrolateral medulla during fatal mevinphos intoxication in the rat.

We evaluated the functional changes in the mitochondrial respiratory chain at the rostral ventrolateral medulla (RVLM), the medullary origin of sympathetic vasomotor tone, in an experimental model of fatal organophosphate poisoning using the insecticide mevinphos (Mev). We also investigated the neuroprotective role of coenzyme Q10 (CoQ10) in this process. Intravenous administration of Mev (1 mg/kg) in Sprague-Dawley rats maintained with propofol elicited an initial hypertension followed by hypotension, accompanied by bradycardia, with death ensuing within 10 min. Enzyme assay revealed a significant depression of the activity of nicotinamide adenine dinucleotide cytochrome c reductase, succinate cytochrome c reductase, and cytochrome c oxidase in the RVLM during this fatal Mev intoxication. ATP production also underwent a significant decrease. Pretreatment by microinjection bilaterally of CoQ10 (4 microg) into the RVLM significantly prevented mortality, antagonized the cardiovascular suppression, and reversed the depressed mitochondrial respiratory enzyme activities, or reduced ATP production in the RVLM induced during Mev intoxication. Our results indicated that dysfunction of mitochondrial respiratory chain and energy production at the RVLM takes place during fatal Mev intoxication. We further demonstrated that CoQ10 provides neuroprotection against Mev-induced cardiovascular depression and fatality through maintenance of activity of the key mitochondrial respiratory enzymes in the RVLM.

Upregulation of FLJ10540, a PI3K-association protein, in rostral ventrolateral medulla impairs brain stem cardiovascular regulation during mevinphos intoxication.

FLJ10540, originally identified as a microtubule-associated protein, induces cell proliferation and migration during tumorigenesis via the formation of FLJ10540-PI3K complex and enhancement of PI3K kinase activity. Interestingly, activation of PI3K/Akt cascade, leading to upregulation of nitric oxide synthase II (NOS II)/peroxynitrite signaling in the rostral ventrolateral medulla (RVLM), the brain stem site that maintains blood pressure and sympathetic vasomotor tone, mediates the impairment of brain stem cardiovascular regulation induced by the pesticide mevinphos. We evaluated the hypothesis that upregulation of FLJ10540 in the RVLM is upstream to this repertoire of signaling cascade that underpins mevinphos-induced circulatory depression. Microinjection bilaterally of mevinphos (10nmol) into the RVLM of anesthetized Sprague-Dawley rats induced a progressive hypotension that was accompanied by an increase (Phase I), followed by a decrease (Phase II) of an experimental index for baroreflex-mediated sympathetic vasomotor tone. There was augmentation in FLJ10540 mRNA in the RVLM or FLJ10540 protein in RVLM neurons, both of which were causally and temporally related to an augmentation of binding between the catalytic subunit (p110) and regulatory subunit (p85) of PI3K, phosphorylation of Akt at Thr308 site, and NOS II, superoxide or peroxynitrite level in the RVLM. Immunoneutralization of FJL10540 in the RVLM significantly antagonized those biochemical changes, and blunted the progressive hypotension and the reduced baroreflex-mediated sympathetic vasomotor tone during mevinphos intoxication. We conclude that upregulation of FLJ10540 in the RVLM elicits impairment of brain stem cardiovascular regulation that underpins circulatory depression during mevinphos intoxication via activation of PI3K/Akt/NOS II/peroxynitrite signaling cascade in the RVLM.

Differential contributions of NOS isoforms in the rostral ventrolateral medulla to cardiovascular responses associated with mevinphos intoxication in the rat.

The organophosphate poison mevinphos (Mev) elicits cardiovascular responses via nitric oxide (NO) produced on activation of M2 muscarinic receptors (M2R) in the rostral ventrolateral medulla (RVLM), where sympathetic vasomotor tone originates. This study further evaluated the contribution of nitric oxide synthase (NOS) isoforms at the RVLM to this process, using adult Sprague-Dawley rats. Bilateral co-microinjection into the RVLM of the selective NOS I inhibitor (250 pmol), 7-nitroindazole or N(omega)-propyl-L-arginine antagonized the initial sympathoexcitatory cardiovascular responses to Mev (10 nmol). Co-administration of a selective NOS II inhibitor, N6-(1-iminoethyl)-L-lysine (250 or 500 pmol) further enhanced these cardiovascular responses and reversed the secondary sympathoinhibitory actions of Mev. A potent NOS III inhibitor, N5-(1-iminoethyl)-L-ornithine (46 or 92 nmol) was ineffective. We also found that M2R co-localized only with NOS I- or NOS II-immunoreactive RVLM neurons. Furthermore, only NOS I or II in the ventrolateral medulla exhibited an elevation in mRNA or protein levels during the sympathoexcitatory phase, with further up-regulated synthesis of NOS II during the sympathoinhibitory phase of Mev intoxication. We conclude that whereas NOS III is not engaged, NO produced by NOS I and II in the RVLM plays, respectively, a sympathoexcitatory and sympathoinhibitory role in the cardiovascular responses during Mev intoxication.

Depression of mitochondrial respiratory enzyme activity in rostral ventrolateral medulla during acute mevinphos intoxication in the rat.

We investigated possible changes in bioenergetics at the rostral ventrolateral medulla (RVLM), a medullary site where sympathetic vasomotor tone originates and where the organophosphate poison mevinphos (Mev) acts to elicit cardiovascular intoxication. In Sprague-Dawley rats maintained under propofol anesthesia, microinjection bilaterally of Mev (10 nmol) into the RVLM induced progressive hypotension that was accompanied by an early augmentation (80-100 min post-Mev; Phase I), followed by a decrease (>100 min post-Mev; Phase II) in the power density of the vasomotor components (0-0.8 Hz) in systemic arterial pressure (SAP) signals. Enzyme assay revealed that local application of Mev into the RVLM also significantly and progressively depressed the activity of NADH cytochrome c reductase (marker for Complexes I and III) and cytochrome c oxidase (marker for Complex IV) in the mitochondrial respiratory chain of the RVLM, but not the heart. On the other hand, the activity of succinate cytochrome c reductase (marker for Complexes II and III) remained unaltered. Both the cardiovascular consequences and depression of mitochondrial respiratory chain enzymes elicited by Mev were significantly antagonized on comicroinjection of atropine (3.5 or 7 nmol) bilaterally into the RVLM. We conclude that Mev adversely effects cardiovascular control by acting as a cholinesterase inhibitor in the RVLM, whose neuronal activity is intimately related to the death process. The resulting accumulation of acetylcholine and prolonged activation of muscarinic receptors in the RVLM is manifested by a selective dysfunction of respiratory enzyme Complexes I and IV in the mitochondrial respiratory chain that underlies cardiovascular toxicity associated with organophosphate poisons such as Mev.

De novo synthesis of ubiquitin carboxyl-terminal hydrolase isozyme l1 in rostral ventrolateral medulla is crucial to survival during mevinphos intoxication.

Ubiquitin carboxyl-terminal hydrolase isozyme L1 (UCH-L1) is a deubiquitinating enzyme that is responsible for making ubiquitin, which is required to target proteins for degradation by the ubiquitin-proteasome pathway in neurons, available. We investigated whether UCH-L1 plays a neuroprotective role at the rostral ventrolateral medulla (RVLM), the origin of sympathetic neurogenic vasomotor tone in the medulla oblongata where the organophosphate insecticide mevinphos (Mev) acts to elicit cardiovascular toxicity. In Sprague-Dawley rats maintained under propofol anesthesia, Mev (960 microg/kg, i.v.) induced a parallel and progressive augmentation in UCH-L1 or ubiquitin expression at the ventrolateral medulla during the course of Mev intoxication. The increase in UCH-L1 level was significantly blunted on pretreatment with bilateral microinjection into the RVLM of a transcription inhibitor, actinomycin D (5 nmol), or a translation inhibitor, cycloheximide (20 nmol). Compared with aCSF or sense oligonucleotide (100 pmol) pretreatment, microinjection of an antisense uch-L1 oligonucleotide (100 pmol) bilaterally into the RVLM significantly increased mortality, reduced the duration of the "pro-life" phase, blunted the increase in ubiquitin expression in ventrolateral medulla, and augmented the induced hypotension in rats that received Mev. These findings suggest that de novo synthesis of UCH-L1, leading to an enhanced disassembly of ubiquitin-protein conjugates in the RVLM, is essential to maintenance of the "pro-life" phase of Mev intoxication via prevention of cardiovascular depression, leading to neuroprotection.

Risks from occupational and dietary exposure to mevinphos.

Mevinphos (trade name, Phosdrin), a category 1 organophosphorus insecticide, has been used mainly as a cleanup pesticide for vegetable crops. A risk assessment for occupational and dietary exposure to mevinphos was initiated because of the high acute toxicity of the compound. Repetitive dosing with mevinphos did not cause any discernible histopathological effects in mice or rats, nor was it oncogenic in either species. The principal toxic effects of mevinphos, both short- and long term, were due to inhibition of cholinesterase activity. Consequently, potential adverse effects from short-term exposures were the primary concern. A human no-observed-effect level (0.025 mg/kg) for cholinergic signs was used as the regulatory basis for calculating margins of safety (MOSs) for potential acute dietary and short-term occupational exposures. Estimates of exposure to mixer/loaders, pilots, and flaggers associated with aerial application of mevinphos were based on passive dosimetry. Because no acceptable exposure studies for work tasks associated with ground application of mevinphos were available, surrogate data based on ground application of oxydemeton-methyl were used. Exposure estimates for field workers and harvesters relied on measured dislodgeable foliar residues of mevinphos and transfer factors generated from studies of other active ingredients. MOSs for mean acute occupational exposure of mixer/loader/applicators associated with ground application and of harvesters working in fruit trees were less than the value conventionally recommended to protect people from the toxic effects of mevinphos. MOSs for the 95th percentile of short-term worker exposure for all mixer/loader work categories associated with mevinphos application were also inadequate. Calculated MOSs for potential acute dietary exposure to measured residue levels of mevinphos were adequate for the various population subgroups. However, 25 of the USEPA tolerances for mevinphos on agricultural commodities were not adequate to protect for the toxic effects of mevinphos from theoretical acute dietary exposure to one or more population subgroups if commodities are consumed with residues at the tolerance level. When the mean short-term occupational exposures were combined with potential acute dietary exposure, the MOSs for mixer/loaders engaged in aerial applications, as well as ground applications, were inadequate to protect people from the toxic effects of mevinphos. As mitigation of the estimated excessive occupational exposures did not appear possible, both California and the USEPA were preparing to cancel registration of the product. However, an agreement was worked out between the manufacturer and the two agencies that ended production for domestic use but allowed existing stocks in the channels of trade to continue to be used for a limited period.

[The effect of parasympatholytics on the therapeutic effectiveness of the oxime HI-6 against organophosphorus compounds (Soman, substance VX, Fosdrin) in mice]. / Vliv vybraných parasympatolytik na terapeutickou úcinnost oximu HI-6 proti organofosforovým slouceninam (Soman, látka VX, Fosdrin) u mysí.

BACKGROUND: Causal antidotal therapy of acute intoxications with organophosphorus compounds involving administration of the parasympatholytic and cholineesterase reactivator (oxime) has not been resolved so far satisfactorily despite knowledge of the basic mechanism of action of these noxious substances. METHODS AND RESULTS: In experiments on mice the therapeutic effect of parasympatholytics atropine, benactyzine and biperidene (Akineton) combined with oxime HI-6 on the toxicity of highly toxic organophosphates soman and substance VX and the organophosphorus insecticide phosdrine was compared as regards their influence on the LD50 of these noxious substances during 24-hour survival of experimental animals. Two levels of antidotes were tested. These findings confirm that the LD50 value of untreated intoxication with all three organophosphorus compounds is most increased by oxime HI-6 combined with benactyzine regardless of the antidote dosage. CONCLUSIONS: Oxime HI-6 is the most effective against highly toxic organophosphates and organophosphorus insecticides when combined with the centrally acting parasympatholytic benactyzine.

[The effect of panpal prophylaxis on acetylcholinesterase activity in the blood, diaphragm and various parts of the brain in rats during treated and untreated poisoning with the organophosphorus insecticide phosdrine]]. / Vliv profylaxe panpalem na aktivitu acetylcholinesterázy v krvi, bránici a ruzných cástech mozku potkana v prubehu nelécené a lécené intoxikace organofosforovým insekticidem fosdrinem.

In experiments on male rats the paper investigated the effect of pharmacological prophylaxis with Panpal (pyridostigmine in combination with benactyzine and trihexyphenidyl) on the activity of acetylcholinesterase in the whole blood, diaphragm, and selected parts of the brain (frontal cortex, pontomedular region, hippocampus, cerebellum) at hour 1 and 3 of untreated and treated (oxime HI-6 with atropine) intoxication with the organophosphorous insecticide phosdrine. Whereas in the CNS Panpal did not produce statistically significant changes in the activity of acetylcholinesterase in the course of untreated and treated phosdrine intoxication, in the blood and diaphragm Panpal markedly intensified phosdrine-induced inhibition of the acetylcholinesterase activity and, in addition, decreased the reactivating effect of the oxime HI-6. The data give evidence of the importance of the combination of the prophylactically used reversible acetylcholinesterase inhibitor pyridostigmine with anticholinergics, which could eliminate the consequences of a pyridostigmine-induced decrease in the activity of the enzyme in the periphery.

[Comparison of the effects of BI-6, a new asymmetric bipyridine oxime, with HI-6 oxime and obidoxime in combination with atropine on soman and fosdrine toxicity in mice]. / Srovnání úcinku nového asymetrického bispyridiniového oximu BI-6 s oximem HI-6 a obidoximem v kombinaci s atropinem na toxicitu somanu a fosdrinu u mysí.

The therapeutic efficacy of the new asymmetric bispyridinium oxime BI-6 against acute toxicity of the highly toxic organophosphate soman and the organophosphorus insecticide fosdrin by means of affecting the LD50 values of these noxiores substances was compared with the effect of the hitherto most perspective oxime HI-6 and the classic obidoxime always in combination with the identical dose of atropine. At the equimolar level the effect of oxime BI-6 against fosdrin completely equals the effects of both oximes HI-6 and obidoxime. The effect of oxime BI-6 against soman is even more marked than the effect of HI-6 but this difference is not statistically significant. On the other hand, at the equi-effective level, the effect of oxime BI-6 against soman is statistically significantly lower than the effect of HI-6, and against fosdrin it is even lower than the effect of both remaining oximes. The effects of the new oxime BI-6 equal, or slightly exceed the therapeutic effect of HI-6 but at the equimolar level only. At the equi-effective level which respects the toxicity of the oxime and is therefore more important for practical use, it is a therapeutically weaker reactivator of acetylcholinesterase than HI-6.

Activation of PI3K/Akt signaling in rostral ventrolateral medulla impairs brain stem cardiovascular regulation that underpins circulatory depression during mevinphos intoxication.

As the most widely used pesticides in the globe, the organophosphate compounds are understandably linked with the highest incidence of suicidal poisoning. Whereas the elicited toxicity is often associated with circulatory depression, the underlying mechanisms require further delineation. Employing the pesticide mevinphos as our experimental tool, we evaluated the hypothesis that transcriptional upregulation of nitric oxide synthase II (NOS II) by NF-κB on activation of the PI3K/Akt cascade in the rostral ventrolateral medulla (RVLM), the brain stem site that maintains blood pressure and sympathetic vasomotor tone, underpins the circulatory depressive effects of organophosphate poisons. Microinjection of mevinphos (10 nmol) bilaterally into the RVLM of anesthetized Sprague-Dawley rats induced a progressive hypotension that was accompanied sequentially by an increase (Phase I) and a decrease (Phase II) of an experimental index for the baroreflex-mediated sympathetic vasomotor tone. There were also progressive augmentations in PI3K or Akt enzyme activity and phosphorylation of p85 or Akt(Thr308) subunit in the RVLM that were causally related to an increase in NF-κB transcription activity and elevation in NOS II or peroxynitrite expression. Loss-of-function manipulations of PI3K or Akt in the RVLM significantly antagonized the reduced baroreflex-mediated sympathetic vasomotor tone and hypotension during Phase II mevinphos intoxication, and blunted the increase in NF-κB/NOS II/peroxynitrite signaling. We conclude that activation of the PI3K/Akt cascade, leading to upregulation of NF-κB/NOS II/peroxynitrite signaling in the RVLM, elicits impairment of brain stem cardiovascular regulation that underpins circulatory depression during mevinphos intoxication.

Transition from oxidative stress to nitrosative stress in rostral ventrolateral medulla underlies fatal intoxication induced by organophosphate mevinphos.

As the most widely used pesticides in the world, fatal incidence of suicidal poisoning by organophosphate compounds is high and is often associated with cardiovascular toxicity. Using the pesticide mevinphos as our tool, we investigated the roles of oxidative stress and nitrosative stress at the rostral ventrolateral medulla (RVLM), the brain stem site that maintains arterial pressure (AP) and sympathetic vasomotor tone, in the cardiovascular depressive effects of organophosphate poisons. Microinjection of mevinphos (10 nmol) into the RVLM of anesthetized Sprague-Dawley rats induced progressive hypotension that was accompanied by an increase (phase I), followed by a decrease (phase II) of an experimental index of baroreflex-mediated sympathetic vasomotor tone, with a fatality rate of 35%. During phase I, there was a preferential upregulation of angiotensin type I receptor (AT1R) messenger RNA (mRNA) and protein that leads to activation of NADPH oxidase (Nox) and increase in superoxide at the RVLM. Pharmacological antagonism of these signals exacerbated fatality and shorted survival time by eliminating baroreflex-mediated sympathetic vasomotor tone, AP, and heart rate. During phase II, there was a progressive upregulation of angiotensin type II receptor (AT2R) mRNA and protein that leads to increase in peroxynitrite in the RVLM, blockade of both sustained brain stem cardiovascular regulation and improved survival. We further found that AT1R and AT2R cross-interacted at transcriptional and signaling levels in the RVLM. We conclude that a transition from AT1R-mediated oxidative stress to AT2R-mediated nitrosative stress in the RVLM underlies the shift from sustained to impaired brain stem cardiovascular regulation that underpins cardiovascular fatality during mevinphos intoxication.