M. Shafiullah

Long‐term effects of streptozotocin‐induced diabetes on the electrocardiogram, physical activity and body temperature in rats

In vivo biotelemetry studies have demonstrated that short‐term streptozotocin (STZ)‐induced diabetes is associated with a reduction in heart rate (HR) and heart rate variability (HRV) and prolongation of QT and QRS intervals. This study investigates the long‐term effects of STZ‐induced diabetes on the electrocardiogram (ECG), physical activity and body temperature. Transmitter devices were surgically implanted in the peritoneal cavity of young adult male Wistar rats. Electrodes from the transmitter were arranged in Einthoven bipolar lead II configuration. ECG, physical activity and body temperature data were continuously recorded with a telemetry system before and following the administration of STZ (60 mg kg−1) for a period of 22 weeks. HR, physical activity and body temperature declined rapidly 3–5 days after the administration of STZ. The effects became conspicuous with time reaching a new steady state approximately 1–2 weeks after STZ treatment. HR at 4 weeks was 268 ± 5 beats min−1 in diabetic rats compared to 347 ± 12 beats min−1 in age‐matched controls. HRV at 4 weeks was also significantly reduced after STZ treatment (18 ± 3 beats min−1) compared to controls (33 ± 3 beats min−1). HR and HRV were not additionally altered in either diabetic rats (266 ± 5 and 20 ± 4 beats min−1) or age‐matched controls (316 ± 6 and 25 ± 4 beats min−1) at 22 weeks. Reduced physical activity and/or body temperature may partly underlie the reductions in HR and HRV. In addition, the increased power spectral low frequency/high frequency ratio from 4 weeks after STZ treatment may indicate an accompanying disturbance in sympathovagal balance.

EFFECTS OF INSULIN TREATMENT ON HEART RHYTHM, BODY TEMPERATURE AND PHYSICAL ACTIVITY IN STREPTOZOTOCIN-INDUCED DIABETIC RAT

1 Streptozotocin (STZ)‐induced diabetic cardiomyopathy is frequently associated with depressed diastolic/systolic function and altered heart rhythm. 2 The effects of insulin treatment on heart rhythm, body temperature and physical activity in STZ‐induced diabetic rats were investigated using biotelemetry techniques. 3 Transmitter devices were surgically implanted in the peritoneal cavity of young adult male Wistar rats. Electrodes from the transmitter were arranged in Einthoven bipolar – Lead II configuration. Electrocardiogram, physical activity and body temperature data were recorded with a telemetry system for 10 days before STZ treatment, for 20 days following administration of STZ (60 mg/kg) and thereafter, for 30 days while rats received daily insulin. 4 Heart rate, physical activity and body temperature declined rapidly 3–5 days after administration of STZ. Pre‐STZ heart rate was 362 ± 7 b.p.m., falling to 266 ± 12 b.p.m. 5–15 days after STZ with significant recovery to 303 ± 14 b.p.m. 10–20 days after commencement of insulin. Pre‐STZ body temperature was 37.5 ± 0.1C, falling to 37.2 ± 0.2C 5–15 days after STZ with significant recovery to 37.5 ± 0.1C 10–20 days after commencement of insulin. Physical activity and heart rate variability were also reduced after STZ but there was no significant recovery during insulin replacement. 5.  Defective autonomic regulation and/or mechanisms of control that are intrinsic to the heart may underlie disturbances in heart rhythm in the STZ‐induced diabetic rat.

Effect of maternal diabetes and ethanol interactions on embryo development in the mouse

The aim of this study was to determine the possible fetal effects of interaction between maternal diabetes and acute doses of alcohol. Pregnant TO mice were made diabetic by a single injection of streptozotocin (STZ) on gestation day (GD) 2. Single dose of 0.003 or 0.03 ml/g body weight of fresh ethanol (25% v/v of absolute alcohol in normal saline) was injected into groups of diabetic and nondiabetic animals on GD 7 or 8. One group of diabetic animals had a daily dose of 6–8 IU of insulin subcutaneously. Fetuses were collected on GD 18. There was a significant increase in the incidence of implantation failure in the diabetes plus ethanol groups and insulin control group. Ethanol injection on GD 7 accentuated diabetes-related embryonic resorption and intrauterine growth retardation (IUGR). This effect was less marked in the diabetic group treated with ethanol on GD 8. Diabetes alone produced a greater incidence of IUGR than ethanol alone. Midfacial hypoplasia and minor anomalies were found more frequently in the combination treatment groups. Holoprosencephaly and thymus hypoplasia observed in diabetic groups were found to be reduced in frequency in the diabetes plus ethanol groups, suggesting an antagonistic type of ethanol--diabetes interaction, stage-dependently. Since severely malformed embryos are known to be resorbed/killed in utero in mice, this reduction might reflect the magnitude of early death of severely malformed embryos. These data suggest that the interaction effects are possibly related to alterations in fundamental developmental processes of early embryos. (Mol Cell Biochem 261: 43–56, 2004)

Toxicokinetics of Aflatoxin in Pregnant Mice

Our objective was to study the toxicokinetics of aflatoxin (AF) in pregnant mice. Aflatoxin B1 (AFB1) was administered intraperitoneally (IP) to groups of pregnant mice in single doses of 20 mg/kg on gestation day (GD) 13 and orally at the same gestational age. Controls received (IP and oral) a proportionate volume of solvent only. Maternal blood was collected at 15, 30, 45, 60, 90, 120, and 150 minutes posttreatment. Their AFB1 contents were determined. Aflatoxin B1 concentrations following maternal exposure to AFB1 were highly correlated with time after exposure. The serum concentrations were predictable and the highest serum levels were seen immediately at 15 minutes in mice given AFs IP and at 30 minutes in those given it orally. The absorption was 5.0 μg/min and elimination was 3.0 μg/min. The toxicokinetics of AFB1 have been delineated. Aflatoxins are easily and rapidly absorbed both from the gastrointestinal tract (GI) tract and through the peritoneum.

Tiapride pre-treatment in acute exposure to paraoxon : Comparison of effects of administration at different points-in-time in rats

AbstractIntroduction: Accidental and suicidal exposures to organophosphorus compounds (OPC) are frequent. The inhibition of esterases by OPC leads to an endogenous ACh poisoning. Recently, the FDA approved, based on animal experiments, for military combat medical use oral pyridostigmine (PSTG) for pre-exposure treatment of soman; the concept is to block the cholinesterase reversibly using the carbamate pyridostigmine in order to deny access to the active site of the enzyme to the irreversible inhibitor (OPC) on subsequent exposure. We have shown previously that tiapride (TIA) is in vitro a weak inhibitor of AChE. We also have shown recently that in rats coadministration of TIA with the organophosphate paraoxon significantly decreases mortality without having an impact on red blood cell cholinesterase (RBC-AChE) activity.Purpose of the study: To establish in a prospective, non-blinded study in a rat model of acute high dose OPC (paraoxon; POX) exposure the ideal point in time for TIA pre-treatment administration and to correlate it with measured TIA plasma levels.Material and Methods: There were six groups of rats in each cycle of the experiment and each group contained six rats. The procedure was repeated twelve times (cycles) (n = 72 for each arm; half male and half female). All substances were applied ip.All groups (1–6) received 1 μMol POX (≈LD75); groups 1–5 also received 50 μMol TIA at different points in time. Group 1 (G1): TIA 120 min before POXGroup 2 (G2): TIA 90 min before POX,Group 3 (G3): TIA 60 min before POX,Group 4 (G4): TIA 30 min before POX,Group 5 (G5): TIA & POX simultaneously,Group 6 (G6): POX only. The animals were monitored for 48 hours and mortality/survival times were recorded at 30 min, 1, 2, 3, 4, 24 and 48 h. AChE activities were determined at 30 min, 24 and 48 h in surviving animals.Statistical analysis was performed on the mortality data, cumulative survival times and enzyme activity data. Mortality data was compared using Kaplan-Meier plots. Cumulative survival times and enzyme activites were compared using the Mann-Whitney rank order test. No Bonferroni correction for multiple comparisons was applied and an α ≤ 0.05 was considered significant.Results: Mortality is statistically significantly reduced by TIA pre-treatment at all points-in-time. Highest protection is achieved if TIA is given 90 to 0 min before OPC exposure. The reduction in mortality is not correlated to TIA plasma levels (Cmax ≈ 120 min post ip-administration). TIA pre-treatment is not affecting AChE activity regardless of the timing of administration.Conclusion: The lack of correlation between TIA plasma levels and degree of mortality reduction as well as the lack of protective effect on enzyme activity seem to indicate that the site of action of TIA is not the blood. While our hypothesis that TIA would protect AChE in a pyridostigmine-like manner (via protection of the enzyme) could not be confirmed, the reduction in mortality with TIA pre-treatment is nevertheless of potential interest. (Mol Cell Biochem xxx: 1–8, 2005)

Effect of intrathecal pralidoxime administration upon survival of rats exposed to the organophosphate paraoxon

The therapeutic results of systemic administration of pralidoxime (2-PAM) in the treatment of poisoning with organophosphate-type cholinesterase inhibitors are disappointing. It has been hypothesized that this is due to poor entry of 2-PAM into the brain. To test if survival rates can be improved by direct administration of 2-PAM into the cerebrospinal fluid (CSF), the effect of intrathecal 2-PAM injections upon mortality after paraoxon intoxication was examined. Eight groups of rats (n=30 each) were examined, all of which received paraoxon (1 micromol=272 microg, 3 micromol=816 microg, or 5 micromol=1.36 mg) intraperitoneally (i.p.). One group received no further treatment; the other groups were given 50 micromol (=8.63 mg) 2-PAM i.p., 5 micromol (=863 microg) 2-PAM intrathecally and pentobarbital/lidocaine in various combinations. Results were compared with the no treatment group and the control groups that did not receive any paraoxon injections, but were given intrathecal injections of saline or 2-PAM. The relative risk of death was estimated by Cox survival analysis. Mortality was lowest after treatment with a combination of both i.p. and intrathecal 2-PAM plus pentobarbital, and with 2-PAM i.p. alone plus pentobarbital. Both treatments were significantly better than 2-PAM i.p. alone (p<or=0.0001). Mortality of intrathecal 2-PAM plus either pentobarbital or lidocaine was in the same order of magnitude as 2-PAM i.p. without pentobarbital/lidocaine, which was significantly (p<or=0.05) lower than that of the no treatment group and of the groups that had only been given pentobarbital or lidocaine. Our results indicate that i.p. injections of 2-PAM insufficiently protect the CNS against the effect of paraoxon. Supplementary injection of pentobarbital, presumably by its anticonvulsive action, has a superior efficacy compared to 2-PAM i.p. alone, irrespective of additional intrathecal 2-PAM administration.

Efficacy of N-Acetylcysteine, Glutathione, and Ascorbic Acid in Acute Toxicity of Paraoxon to Wistar Rats: Survival Study.

There are a great number of reports with assertions that oxidative stress is produced by organophosphorus compound (OPC) poisoning and is a cofactor of mortality and morbidity in OPC toxicity. In addition, antioxidants have been suggested as adjuncts to standard therapy. However, there is no substantial evidence for the benefit of the use of antioxidants in survival after acute intoxication of OPCs. The present study was conducted to assess the effectiveness of three non-enzymatic antioxidants (NEAOs), N-acetylcysteine (NAC), glutathione (GSH), and ascorbic acid (AA), in acute intoxication of adult male Wister rats with paraoxon. The efficacy of the antioxidants was estimated as both a pretreatment and a concurrent application along with the standard oxime, pralidoxime (2-PAM). Relative risk of death after 48 hours of application was estimated by Cox regression analysis. The results revealed no benefit of either tested NEAO to the improvement in survival of experimental rats. The application of these antioxidants was found to be deleterious when administered along with pralidoxime compared to the treatment with pralidoxime alone. It has been concluded that the tested non-enzymatic antioxidants are not useful in acute toxicity for improving survival rates. However, the individual toxic dynamics of diversified OPCs should not be overlooked and further studies with different OPCs are suggested.

Effects of Pioglitazone on the Electrocardiogram in the Goto-Kakizaki Type 2 Diabetic Rat Heart

Cardiovascular complications are the major cause of morbidity and mortality in diabetic patients. Pioglitazone (PIO) is used for the treatment of type 2 diabetes mellitus and there is some evidence that it may improve ventricular function in diabetic patients. The effects of PIO on electrical conduction in the Goto-Kakizaki (GK) type 2 diabetic rat heart have been investigated. Transmitter devices were surgically implanted in GK and control rats aged 7 months. PIO at concentrations (2.5 to 20 mg/kg) was added to the drinking water. Animals received each concentration of PIO for 2 weeks. Bodyweight and blood glucose were measured periodically. The electrocardiogram was recorded throughout the study. Blood glucose was higher in GK rats compared to controls and was progressively reduced in GK rats treated with increasing concentrations of PIO. A modest reduction in heart rate was associated with a significant increase in PQ interval in GK rats compared to controls. Heart rate and PQ interval were not additionally altered by PIO. QRS complex and QT interval were not altered in GK compared to control or by PIO. In conclusion daily oral doses of PIO in the range 2.5 to 20 mg/ kg during a period of 14 weeks appeared to have little effect on electrical conduction in GK and control rat heart.