Alloxan: An unpredictable drug for diabetes induction?

Monika Misra, Umme Aiman 
 Department of Pharmacology, Jawaharlal Nehru Medical College, Aligarh Muslim University, Aligarh, India

Correspondence Address:
Monika Misra
Department of Pharmacology, Jawaharlal Nehru Medical College, Aligarh Muslim University, Aligarh
India

Full Text

Sir,

We read with interest the article titled "Anomalies in alloxan-induced diabetic model: It is better to standardize it first" by Jain et al., [1] wherein the authors discussed the inconsistencies associated with alloxan-induced experimental diabetes in animals. We used alloxan to induce diabetes in rats and rabbits for screening the antidiabetic activity of test compounds. However, we experienced an inconsistent and unpredictable response with alloxan, similar to that discussed by Jain et al.

Healthy adult albino rats (150-200 g) and rabbits (1.5-2.5 kg) were used in the study. Alloxan monohydrate (Sigma Aldrich, St. Louis, MO, USA; stored at 4°C) was dissolved in normal saline at room temperature and was injected by intraperitoneal and intravenous (i.v.) routes, respectively in overnight fasted rats and rabbits. Rats were divided into three groups: 12 rats received 150 mg/kg, 6 rats received 160 mg/ kg, and 10 rats received 170 mg/kg dose of alloxan. Alloxan (150 mg/kg as a 5% solution in normal saline) was given to rabbits via marginal ear vein. Rabbits were divided into two groups: 10 rabbits received alloxan by slow i.v. injection over 15 minutes and 10 rabbits received alloxan by rapid i.v. injection. The animals were then kept for the next 24 hours on 10% glucose to prevent hypoglycemia. After 72 hours, fasting blood glucose (FBG) was determined by using Accuchek glucometer strips (Roche Diagnostics). Animals with FBG >200 mg/dl were considered to have developed experimental diabetes. Thereafter, FBG was recorded at days 3, 7, 14, 21, 28, and 35 to determine fluctuations in FBG or autoreversal of diabetes. The Institutional Animal Ethics Committee approved the experimental protocol.

Alloxan induced a dose-dependent mortality in 25%, 33.33%, and 40% of rats receiving 150 mg/kg, 160 mg/kg, and 170 mg/ kg of alloxan within 15 days of injection, respectively. In rabbits, a very high mortality of about 80% was seen with rapid i.v. injection of alloxan compared to 30% with slow i.v. injection. Alloxan induced diabetes in 33.33% of rats receiving either 150 mg/kg or 160 mg/kg of alloxan and 60% of rats receiving 170 mg/kg dose. All surviving rabbits given alloxan via rapid i.v. injection developed diabetes compared to 60% of surviving rabbits given slow i.v. injection. In diabetic rats and rabbits, random fluctuations in FBG with blood sugar returning to the nondiabetic range at various time periods during the study duration was observed. These fluctuations had no discernible pattern. These were noted with both low and high doses of alloxan in rats. Stable diabetes for 35 days was produced in 20% of rats receiving 170 mg/kg of alloxan and rabbits given slow i.v. injection. The rats that were nondiabetic after initial dose of alloxan showed no impairments in oral glucose tolerance test and remained resistant to two subsequent doses of alloxan injected at 1 week interval.

These results indicate that the susceptibility to diabetogenic and toxic effects of alloxan differs among animals of the same species. Such unpredictable responses with alloxan have also been reported by authors in the past. [2] As observed in our study, a high mortality (30-60%) of experimental animals with alloxan has been reported in previous studies. [2] This mortality is attributed either due to initial hypoglycemic shock or emergence of diabetic complications or direct kidney tubular cell toxicity. [3] A wide range of fluctuations in FBG returning to the nondiabetic range is a chief drawback of using alloxan for experimental diabetes induction, as this would create an illusion of positive antidiabetic activity with the test drug. Autoreversal from frank diabetic state has been reported with suboptimal doses (90-140 mg/kg) of alloxan, which is due to regeneration of pancreatic beta cells. [1] However, in our study, autoreversal from diabetic state was seen with relatively high dose of alloxan (170 mg/kg) in one rat. The high degree of resistance to alloxan in rats suggests genetic differences in the constitutive ability to dissipate reactive oxygen species responsible for diabetogenic effect of alloxan. [4] The induction of diabetes depends upon speed of the i.v. injection at a given dose level, with slower injection rates causing less effect [5] Similar to these reports, we found a high success rate in diabetes induction by rapid i.v. injection in rabbits. This, however, was also associated with a significantly increased mortality.

The results of this study suggest the inconsistencies associated with use of alloxan in induction and maintenance of a stable diabetic state over a period of 1 month in both rats and rabbits. Thus, with these fluctuations and unpredictable responses at any dose level, we were unable to derive any hypothesis about the hypoglycemic property of test compounds. We, therefore, agree with the viewpoint of Jain DK et al. that alloxan is a doubtful drug as regards the induction of experimental diabetes. This, however, remains contradictory to many studies where alloxan has been successfully used for the same.

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