Antonio Monroy-Noyola

d-penicillamine and prussian blue as antidotes against thallium intoxication in rats

D-penicillamine (DP) and prussian blue (PB), given alone and in combination, were evaluated in rats as treatments against acute thallotoxicosis. Animals were poisoned by intraperitoneal (i.p.) injection of thallium(I) acetate at different doses (16, 30, 40, 50 and 70 mg/kg). Later (24 h), treatments were administered until day 5, as follows: D-penicillamine (DP), 25 mg/kg, i.p. route, twice daily; prussian blue (PB), 50 mg/kg, oral route, twice daily. LD50 values were estimated for each treatment with the following results: control, 32 mg/kg; DP, 27 mg/kg; PB, 42 mg/kg; PB + DP, 64 mg/kg. Thallium content was analyzed in six body organs and eight brain regions after treatments. PB administration induced significant elimination of thallium from all tissues. DP treatment diminished thallium content in body organs, but increased it in brain regions, indicating a redistributive effect of DP. DP + PB treatment decreased thallium content in all body organs and brain regions. Renal thallium content in the DP + PB group was significantly lower than that of PB alone group, suggesting accelerated urinary excretion of thallium as a result of DP action. Results indicate that DP administered alone may be dangerous because of its redistributive effect, but given in combination with PB may be useful as treatment against thallium poisoning.

Copper reduces striatal protein nitration and tyrosine hydroxylase inactivation induced by MPP+ in rats

Striatal administration of 1-methyl-4-phenylpyridinium (MPP(+)), the active metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), causes nigrostriatal dopaminergic pathway damage similar to that observed in Parkinsons disease. Copper acts as a prosthetic group of several antioxidant enzymes and recent data show that copper attenuated MPP(+)-evoked neurotoxicity. We evaluated the effect of copper (as a supplement) upon proteins nitration (60 kDa) and tyrosine hydroxylase (TH) inactivation induced by MPP(+) (10 microg/8 microL) injection into the rat striatum. Copper pretreatment (10 micromol/kg i.p.) prevented both MPP(+)-induced proteins nitration and TH inactivation. Copper treatment also prevented the dopamine-depleting effect of MPP(+) injection. Those results were accompanied by a significant reduction of enzymatic activity of the constitutive nitric oxide synthase (cNOS), whereas, the protein levels of the three isoforms of NOS remained unchanged. Results indicate that the effect of copper against MPP(+)-induced proteins nitration and TH inactivation in the striatum of rat may be mediated by a reduction of cNOS activity.

EGb761 Pretreatment Reduces Monoamine Oxidase Activity in Mouse Corpus Striatum During 1-Methyl-4-Phenylpyridinium Neurotoxicity

EGb761 produces reversible inhibition of both monoamine oxidase (MAO) isoforms in the central nervous system. 1-methyl-4-phenylpyridinium (MPP+) neurotoxicity is prevented by treatment with the MAO inhibitor pargyline. We investigated EGb761s effect on striatal MAO activity during MPP+ neurotoxicity. C-57 black mice were pretreated with EGb761 (10 mg/kg) daily for 17 days followed by administration of MPP (0.72 mg/kg). MPP+ enhanced striatal MAO (30%) activity at 6 h, and EGb761 prevented this effect. MAO-B activity in striatum was enhanced (70%) 6 h after MPP+ administration and was reduced to almost normal levels in EGb761 + MPP+ group compared to MPP+ group. Pretreatment with EGb761 partially prevented (32%) the striatal dopamine-depleting effect of MPP+ and prevented the reduction in striatal tyrosine hydroxylase activity (100%). Results suggest that EGb761 supplements may be effective in reducing MAO activity as well as enhancement in dopamine metabolism, thereby preventing MPP+-neurotoxicity.

Combined D-penicillamine and prussian blue as antidotal treatment against thallotoxicosis in rats: evaluation of cerebellar lesions

Rats were treated with a single dose of thallium acetate (32 mg/kg i.p.) and the antidotal effect of D-penicillamine and prussian blue given alone or in combination was assessed by means of evaluation of the thallium-induced cerebellar histological lesions. After thallium poisoning (24 h), antidotes were administered for 4 days as follows: D-penicillamine (DP) 25 mg/kg, i.p. twice daily; prussian blue (PB), 50 mg/kg p.o., twice daily. Mortality among the treatment groups was as follows: control, 87.5%; DP, 100%; PB, 56.25%; DP+PB, 25%. Three days after these treatments, rats treated with the combination DP+PB presented a significantly lower number of altered Purkinje cells in cerebellum as compared with those of the thallium alone treated animals, indicating adequate protection by this antidote treatment against thallium neurotoxicity. Prussian blue protected against thallium-induced neurotoxicity to a lesser extent as compared with the effects obtained by the DP+PB protection. DP did not protect against thallium-induced alterations of Purkinje cells. These results confirm the efficacy of the combined antidotal treatment of DP and PB against thallium toxicity in rats, and support the possible application in human cases of thallotoxicosis.

Metallothionein-II Inhibits Lipid Peroxidation and Improves Functional Recovery after Transient Brain Ischemia and Reperfusion in Rats

After transient cerebral ischemia and reperfusion (I/R), damaging mechanisms, such as excitotoxicity and oxidative stress, lead to irreversible neurological deficits. The induction of metallothionein-II (MT-II) protein is an endogenous mechanism after I/R. Our aim was to evaluate the neuroprotective effect of MT-II after I/R in rats. Male Wistar rats were transiently occluded at the middle cerebral artery for 2 h, followed by reperfusion. Rats received either MT (10 μg per rat i.p.) or vehicle after ischemia. Lipid peroxidation (LP) was measured 22 h after reperfusion in frontal cortex and hippocampus; also, neurological deficit was evaluated after ischemia, using the Longa scoring scale. Infarction area was analyzed 72 hours after ischemia. Results showed increased LP in frontal cortex (30.7%) and hippocampus (26.4%), as compared to control group; this effect was fully reversed by MT treatment. Likewise, we also observed a diminished neurological deficit assessed by the Longa scale in those animals treated with MT compared to control group values. The MT-treated group showed a significant (P < 0.05) reduction of 39.9% in the infarction area, only at the level of hippocampus, as compared to control group. Results suggest that MT-II may be a novel neuroprotective treatment to prevent ischemia injury.

Induction of ferroxidase enzymatic activity by copper reduces MPP + -evoked neurotoxicity in rats

Parkinsons disease (PD) is a neurodegenerative disorder characterized by decreased dopamine, intracellular inclusions (Lewy bodies) and brain iron deposits. PD has also been related with reduced ferroxidase activity, diminished antioxidant defenses and lipid peroxidation. Striatal injection of 1-methyl-4-phenylpyridinium (MPP(+)) into rodents reproduces the major biochemical characteristics of PD, including oxidative stress. Copper (Cu) plays an important role as prosthetic group of several proteins involved in iron metabolism and antioxidant responses, such as ceruloplasmin (Cp). In the present study, intraperitoneal CuSO4 injection (10μmol/kg) produced an insignificant increase of Cu content in striatum and midbrain (17.5% and 7%, respectively). After 10 and 11h, Cu induced 6- and 4-fold increase Cp mRNA in midbrain and striatum, respectively. Cu-supplement also produced a time-dependent increase ferroxidase activity in striatal tissue, reaching a maximum 16h after Cu treatment in midbrain; while, ferrous iron content diminished 18% in striatum and 8% in midbrain. In regard the PD model, we found that MPP(+) (10μg/8μL, intrastriatal), induced a significant (P<0.05) reduction of striatal ferroxidase activity; this effect was reverted by Cu pre-treatment 16h before MPP(+). Likewise, Cu-supplement prevented lipid fluorescent products formation in striatum, evaluated (P<0.01) 6h after MPP(+). In the long term, apomorphine-evoked circling behavior was evaluated 6 days after MPP(+) injury; Cu pre-treatment significantly reduced (P<0.05) the apomorphine-induced ipsilateral turns in MPP(+)-lesioned rats. These results suggest that Cu-induced expression of Cp could be an interesting scope against the deleterious effects of iron deposits in PD.

Additive effect of dl-penicillamine plus Prussian blue for the antidotal treatment of thallotoxicosis in rats

DL-penicillamine (DL-P) and Prussian blue (PB) given alone or in combination were tested as possible treatments against acute thallium toxicity. Rats were intoxicated by i.p. injection of thallium (I) acetate at LD(50) (32 mg/kg). A day later, pharmacological treatment was administered until day 4 as follows: (1) vehicles, (2) PB 50mg/kg, by oral route, twice a day, (3) DL-P 25mg/kg i.p. route, twice daily and (4) PB+DL-P. The Estimated Probability Survival (EPS) was recorded during the experiment for each treatment. DL-P alone did not show a significant effect on survival. However, when it was used in combination with PB, it increased the survival significantly (EPS=0.8, P<0.05) as compared to the control group (EPS=0.4). In a different experiment, using 16 mg/kg of Thallium I acetate, the metal levels were analyzed in blood, body organs and brain regions after treatments. DL-P given alone decreased slightly the thallium content in blood, organs and brain. Meanwhile, its administration in combination with PB diminished the thallium levels significantly (P<0.05) in the majority of tissues, at levels lower than those achieved in the PB group. Those results indicate that DL-P administered alone did not prevent the mortality nor accumulation of the metal in body tissues. Its combination with PB could be considered an alternative antidotal treatment in thallium toxicity, because this chelating agent given alone did not cause thallium redistribution to the brain. When given in combination with PB it has an additive effect in the treatment of acute thallotoxicosis.

The neuroprotective effect of lovastatin on MPP(+)-induced neurotoxicity is not mediated by PON2.

Parkinsons disease (PD) is a neurodegenerative disorder characterized by loss of the pigmented dopaminergic neurons in the substantia nigra pars compacta with subsequent striatal dopamine (DA) deficiency and increased lipid peroxidation. The etiology of the disease is still unclear and it is thought that PD may be caused by a combination of genetic and environmental factors. In the search of new pharmacological options, statins have been recognized for their potential application to treat PD, due to their antioxidant effect. The aim of this work is to contribute in the characterization of the neuroprotective effect of lovastatin in a model of PD induced by 1-methyl-4-phenylpyridinium (MPP(+)). Male Wistar rats (200-250 g) were randomly allocated into 4 groups and administered for 7 days with different pharmacological treatments. Lovastatin administration (5 mg/kg) diminished 40% of the apomorphine-induced circling behavior, prevented the striatal DA depletion and lipid peroxides formation by MPP(+) intrastriatal injection, as compared to the group of animals treated only with MPP(+). Lovastatin produced no change in paraoxonase-2 (PON2) activity. It is evident that lovastatin conferred neuroprotection against MPP(+)-induced protection but this effect was not associated with the induction of PON2 in the rat striatum.

Screening for marijuana and cocaine abuse by immunoanalysis and gas chromatography.

Drug abuse among college students is characterized by lower academic performance and long‐term negative consequences. Screening to detect students at high risk of consuming drugs is of primary importance to insure early identification and appropriate levels of care. As a result, this study aimed to determine the current or past use of drug abuse through a questionnaire applied to a student population at the Universidad Autónoma del Estado de Morelos. The results were confirmed by immunoanalysis and gas chromatography of urine. We interviewed 181 students aged 15 to 21 (gender was not considered in this study), and urine samples were collected for analytical analysis. For detection of metabolites Delta9‐THCA‐A and benzoylecgonine from marijuana and cocaine, respectively, a homogenous enzymatic inmmunoanalysis was used; subsequent samples were analyzed by a mass spectrometer with quadrupole detector. Seven samples of the total (181) did not completely fit the inclusion criteria and were eliminated. The results showed 0.50% and 1.16% positive samples for benzoylecgonine and Delta9‐THCA‐A, respectively. These results are not different from those of the National Questionnaire on Addiction. We can establish a program for detecting drug consumption in our students. This kind of study is important in order to implement programs that can help us to decrease the abuse of drugs in our college population.

Comparative hydrolysis of O-hexyl O-2,5-dichlorophenyl phosphoramidate and paraoxon in different tissues of vertebrates

The Ca2+-dependent and EDTA-resistant hydrolysis of O-hexyl O-2,5-dichlorophenyl phosphoramidate (HDCP) and paraoxon was studied in serum and subcellular fractions of liver, kidney and brain of hen, rat and rabbit. HDCP was the best substrate among all the tissues studied, except that of rabbit serum which showed the highest Ca2+-dependent paraoxon hydrolysing activity (paraoxonase). High HDCP hydrolysing activity (HDCPase) was detected in the brain tissue of the three species studied, whereas low or no paraoxonase was found. The HDCPase/paraoxonase ratio of Ca2+-dependent hydrolysing activities ranged from 0.5 to 83 for tissues of the same species. EDTA-resistant HDCPase activity was more than 50% of the total activities in hen tissues, with an almost undetectable Ca2+-dependent paraoxonase activity in most organs. The same response was observed in rat tissues, except for serum where the Ca2+-dependent HDCPase and paraoxonase activities were higher (70 and 25% of total activities, respectively). EDTA-resistant HDCPase and paraoxonase activities represented less than 25% of all activities in rabbit tissues. Paraoxon has traditionally been the substrate for measuring organophosphorus hydrolysing activities. However, HDCP could be a good substrate in addtion to paraoxon for monitoring other phosphotriesterases in biological tissues.